U.S. patent application number 10/372717 was filed with the patent office on 2004-01-22 for biopolar trans carotenoid salts and their uses.
Invention is credited to Gainer, John L., Grabiak, Raymond C..
Application Number | 20040014725 10/372717 |
Document ID | / |
Family ID | 27765977 |
Filed Date | 2004-01-22 |
United States Patent
Application |
20040014725 |
Kind Code |
A1 |
Gainer, John L. ; et
al. |
January 22, 2004 |
Biopolar trans carotenoid salts and their uses
Abstract
The invention relates to trans carotenoid salt compounds,
methods for making them, methods for solubilizing them and uses
thereof. These compounds are useful in improving diffusivity of
oxygen between red blood cells and body tissues in mammals
including humans.
Inventors: |
Gainer, John L.;
(Charlottesville, VA) ; Grabiak, Raymond C.;
(Maryland Heights, MO) |
Correspondence
Address: |
NIXON & VANDERHYE, PC
1100 N GLEBE ROAD
8TH FLOOR
ARLINGTON
VA
22201-4714
US
|
Family ID: |
27765977 |
Appl. No.: |
10/372717 |
Filed: |
February 25, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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60358718 |
Feb 25, 2002 |
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Current U.S.
Class: |
514/102 ;
514/553; 514/559; 554/221; 558/152; 558/24; 562/20 |
Current CPC
Class: |
A61P 7/00 20180101; A61P
3/00 20180101; C07C 57/13 20130101; A61P 9/12 20180101; A61P 11/00
20180101; A61P 15/14 20180101; C07C 69/602 20130101; A61P 9/00
20180101; A61P 7/02 20180101; A61P 25/00 20180101; A61P 35/00
20180101; A61P 15/00 20180101; A61P 39/06 20180101; A61P 9/10
20180101; A61P 7/10 20180101; A61P 3/10 20180101; A61P 11/06
20180101; C07F 9/113 20130101; A61P 25/28 20180101; C07C 305/14
20130101 |
Class at
Publication: |
514/102 ;
514/553; 514/559; 554/221; 562/20; 558/24; 558/152 |
International
Class: |
A61K 031/663; A61K
031/203; C07F 009/28; A61K 031/185 |
Claims
What is claimed is:
1. A compound having the structure: YZ-TCRO-ZY where: Y=a cation
Z=polar group which is associated with the cation, and TCRO=trans
carotenoid skeleton wherein said compound is not TSC.
2. A compound as in claim 1 wherein Y is a monovalent metal ion
selected from the group consisting of Na.sup.+, K.sup.+, Li.sup.+,
or an organic cation selected from the group consisting of
R.sub.4N.sup.+, R.sub.3S.sup.+, where R is H, or C.sub.nH.sub.2n+1
where n is 1-10.
3. A compound as in claim 1 wherein Z is selected from the group
consisting of a carboxyl (COO.sup.-) group, a sulfate group
(OSO.sub.3.sup.-) or a monophosphate group (OPO.sub.3.sup.-),
(OP(OH)O.sub.2.sup.-), a diphosphate group, triphosphate or
combinations thereof.
4. A compound as in claim 1 wherein TCRO is conjugated
carbon-carbon double and single bonds containing carbon atoms
wherein the 4 single bonds which surround a carbon-carbon double
bond all lie in the same plane and said compound is linear.
5. A compound as in claim 1 wherein TCRO is 28where X which can be
the same or different, is H, a linear or branched group having 10
or less carbons optionally containing a halogen, or a halogen.
6. A compound as in claim 1 wherein TCRO is 29where X which can be
the same or different, is H, a linear or branched group having 10
or less carbons optionally containing a halogen, or a halogen
7. A compound as in claim 1 wherein TCRO is 30where X which can be
the same or different, is H, a linear or branched group having 10
or less carbons optionally containing a halogen, or a halogen.
8. A compound as in claim 1 wherein TCRO is 31where X which can be
the same or different, is H, a linear or branched group having 10
or less carbons optionally containing a halogen, or a halogen.
9. A method of solubilizing a BTCS having the structure YZ-TCRO-ZY
where: Y=a cation Z=a polar group which is associated with the
cation, and TCRO=trans carotenoid skeleton comprising the steps of:
a) preparing a dilute solution of sodium carbonate or sodium
bicarbonate, b) adding said dilute solution to deionized water to
raise the pH to 7 or above, c) adding a BTCS to the solution of
step b).
10. A method of solubilizing a BTCS having the structure YZ-TCRO-ZY
where: Y=a cation Z=a polar group which is associated with the
cation, and TCRO=trans carotenoid skeleton comprising the steps of:
a) adding a BTCS to a saline solution, b) removing undissolved
material.
11. A method of solubilizing a BTCS having the structure YZ-TCRO-ZY
where: Y=a cation Z=a polar group which is associated with the
cation, and, TCRO=trans carotenoid skeleton. comprising the steps
of: a) adding a base to water to make a basic solution, c) adding a
BTCS to said solution.
12. A method of solubilizing a BTCS having the structure YZ-TCRO-ZY
where: Y=a cation Z=a polar group which is associated with the
cation, and TCRO=trans carotenoid skeleton comprising the steps of:
a) preparing deionized water, b) adding a BTCS to the solution of
step a).
13. A method as in claim 9,10,11 or 12 wherein said compound is
trans sodium crocetinate.
14. A method of increasing the diffusivity of oxygen in a mammal
comprising administering to a mammal a therapeutically effective
amount of a compound having the formula: YZ-TCRO-ZY where: Y=a
cation Z=a polar group which is associated with the cation, and
TCRO=trans carotenoid skeleton, wherein said compound is not
TSC.
15. A method as in claim 14 wherein said administration is by
inhalation.
16. A method of treating respiratory disease comprising
administering to a mammal in need of treatment a therapeutically
effective amount of a compound having the formula: YZ-TCRO-ZY
where: Y=a cation Z=a polar group which is associated with the
cation, and TCRO=trans carotenoid skeleton, wherein said compound
is not TSC.
17. A method of treating emphysema comprising administering to a
mammal in need of treatment a therapeutically effective amount of a
compound having the formula YZ-TCRO-ZY where: Y=a cation Z=a polar
group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
18. A method of treating hemorrhagic shock comprising administering
to a mammal in need of treatment a therapeutically effective amount
of a compound having the formula YZ-TCRO-ZY where: Y=a cation Z=a
polar group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
19. A method of treating cardiovascular disease comprising
administering to a mammal in need of treatment a therapeutically
effective amount of a compound having the formula YZ-TCRO-ZY where:
Y=a cation Z=a polar group which is associated with the cation, and
TCRO=trans carotenoid skeleton, wherein said compound is not
TSC.
20. A method of treating atherosclerosis comprising administering
to a mammal in need of treatment a therapeutically effective amount
of a compound having the formula YZ-TCRO-ZY where: Y=a cation Z=a
polar group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
21. A method of treating asthma comprising administering to a
mammal in need of treatment a therapeutically effective amount of a
compound having the formula YZ-TCRO-ZY where: Y=a cation Z=a polar
group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
22. A method of treating spinal cord injuries comprising
administering to a mammal in need of treatment a therapeutically
effective amount of a compound having the formula YZ-TCRO-ZY where:
Y a cation Z=a polar group which is associated with the cation, and
TCRO=trans carotenoid skeleton, wherein said compound is not
TSC.
23. A method of treating cerebral edema comprising administering to
a mammal in need of treatment a therapeutically effective amount of
a compound having the formula YZ-TCRO-ZY where: Y=a cation Z=a
polar group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
24. A method of treating papillomas comprising administering to a
mammal in need of treatment a therapeutically effective amount of a
compound having the formula YZ-TCRO-ZY where: Y=a cation Z=a polar
group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
25. A method of treating hypoxia comprising administering to a
mammal in need of treatment a therapeutically effective amount of a
compound having the formula YZ-TCRO-ZY where: Y=a cation Z=a polar
group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
26. A method of synthesizing a BTCS compound having the formula
YZ-TCRO-ZY where: Y=a cation Z=a polar group which is associated
with the cation, and TCRO=trans carotenoid skeleton, comprising the
steps of: a) coupling a symmetrical dialdehyde containing
conjugated carbon-carbon double bonds with a triphenylphosphorane,
b) saponifying the product of step a).
27. A method as in claim 26 wherein the coupling of step a) is made
using [3-carbomethoxy-2-buten-1-ylidene]triphenylphosphorane.
28. A method as in claim 26 wherein the product of step a) is
saponified using a solution of NaOH and methanol.
29. A method as in claim 26 wherein after step a) is the step of
isolating the desired product of the coupling reaction.
30. A method of saponifying a symmetrical diester containing
conjugated carbon-carbon double bonds to form a BTCS, comprising
the steps of: a) solubilizing the symmetrical diester containing
conjugated carbon-carbon double bonds with a compound selected from
the group consisting of methanol, ethanol, propanol and
isopropanol, and b) mixing the solution of step a) with a base.
31. A method as in claim 30 wherein the base is selected from the
group consisting of NaOH, KOH, and LiOH.
32. A method as in claim 30 wherein the diester is saponified using
methanol and NaOH.
33. A BTCS compound synthesized according to claim 26.
34. A BTCS composition wherein absorbency of the highest peak which
occurs in the visible wave length range divided by the absorbency
of the peak which occurs in the UV wave length range is greater
than 8.5.
35. A TSC composition wherein absorbency of the highest peak which
occurs in the visible wave length range divided by the absorbency
of the peak which occurs in the UV wave length range is greater
than 8.5.
36. A method of increasing the diffusivity of oxygen in a mammal
comprising administering to a mammal a therapeutically effective
amount of BTCS wherein absorbency of the highest peak which occurs
in the visible wave length range divided by the absorbency of the
peak which occurs in the UV wave length range is greater than
8.5.
37. A method of treating emphysema comprising administering to a
mammal in need of treatment a therapeutically effective amount of
BTCS wherein absorbency of the highest peak which occurs in the
visible wave length range divided by the absorbency of the peak
which occurs in the UV wave length range is greater than 8.5.
38. A method of treating hemorrhagic shock comprising administering
to a mammal in need of treatment a therapeutically effective amount
of BTCS wherein absorbency of the highest peak which occurs in the
visible wave length range divided by the absorbency of the peak
which occurs in the UV wave length range is greater than 8.5.
39. A method as in claim 36, 37 or 38 wherein the BTCS is TSC.
40. A method of increasing the diffusivity of oxygen in a mammal
comprising administering to a mammal by inhalation a
therapeutically effective amount of TSC.
41. An inhaler containing a BTCS compound having the structure:
YZ-TCRO-ZY where: Y=a cation Z=polar group which is associated with
the cation, and TCRO=trans carotenoid skeleton.
42. An inhaler as in claim 40 wherein said BTCS compound is
TSC.
43. A method of converting an isomeric mixture of olefinic
dialdehydes into the all trans aldehyde comprising isomerizing said
isomeric mixture of dialdehydes with a sulfinic acid in a
solvent.
44. A method as in claim 43 wherein said sulfinic acid has the
formula RSO2H where R is C1 through C10 straight or branched alkyl
group or an aryl group.
45. A method as in claim 43 where the solvent is selected from the
group consisting of 1,4-dioxane, tetrahydrofuran or dialkyl ether
wherein the alkyl group is a C1 through C10 straight or branched
alkyl group.
46. A method as in claim wherein said sulfinic acid is
para-toluenesulfinic acid and said solvent is 1,4-dioxane.
47. A method as in claim 43 wherein said olefinic dialdehyde is
2,7-dimethyl-2,4,6-ocatrienedial.
48. A method as in claim 43 wherein said olefinic dialdehyde is
2,7-dimethyl-2,4,6-ocatrienedial, said sulfinic acid is
para-toluenesulfinic acid and said solvent is 1,4-dioxane.
49. A method of treating ischemia comprising administering to a
mammal in need of treatment a therapeutically effective amount of a
compound having the formula: YZ-TCRO-ZY where: Y a cation Z=a polar
group which is associated with the cation, and TCRO=trans
carotenoid skeleton, wherein said compound is not TSC.
50. A method of treating traumatic brain injury comprising
administering to a mammal in need of treatment a therapeutically
effective amount of a compound having the formula YZ-TCRO-ZY where:
Y=a cation Z=a polar group which is associated with the cation, and
TCRO=trans carotenoid skeleton, wherein said compound is not
TSC.
51. A method of enhancing performance of a mammal comprising
administering to said mammal a therapeutically effective amount of
a compound having the formula: YZ-TCRO-ZY where: Y=a cation Z=a
polar group which is associated with the cation, and TCRO=trans
carotenoid skeleton.
52. A method of treating complications of diabetes comprising
administering to a mammal in need of treatment a therapeutically
effective amount of a compound having the formula YZ-TCRO-ZY where:
Y=a cation Z=a polar group which is associated with the cation, and
TCRO=trans carotenoid skeleton.
53. A method of treating Alzheimer's disease comprising
administering to a mammal in need of treatment a therapeutically
effective amount of a compound having the formula YZ-TCRO-ZY where:
Y=a cation Z=a polar group which is associated with the cation, and
TCRO=trans carotenoid skeleton.
54. A method of treating ischemia in a mammal comprising
administering to a mammal a therapeutically effective amount of
BTCS wherein absorbency of the highest peak which occurs in the
visible wave length range divided by the absorbency of the peak
which occurs in the UV wave length range is greater than 8.5.
55. A method of treating traumatic brain injury comprising
administering to a mammal in need of treatment a therapeutically
effective amount of BTCS wherein absorbency of the highest peak
which occurs in the visible wave length range divided by the
absorbency of the peak which occurs in the UV wave length range is
greater than 8.5.
56. A method of enhancing performance comprising administering to a
mammal an effective amount of BTCS wherein absorbency of the
highest peak which occurs in the visible wave length range divided
by the absorbency of the peak which occurs in the UV wave length
range is greater than 8.5.
57. A method of treating diabetes comprising administering to a
mammal in need of treatment a therapeutically effective amount of
BTCS wherein absorbency of the highest peak which occurs in the
visible wave length range divided by the absorbency of the peak
which occurs in the UV wave length range is greater than 8.5.
58. A method of treating Alzheimer's disease comprising
administering to a mammal in need of treatment a therapeutically
effective amount of BTCS wherein absorbency of the highest peak
which occurs in the visible wave length range divided by the
absorbency of the peak which occurs in the UV wave length range is
greater than 8.5.
59. A method as in claim 54, 55,56,57 or 58 wherein the BTCS is
TSC.
Description
[0001] The invention relates to bipolar trans carotenoid salt
compounds, methods of solubilizing them, methods for making them,
and methods of using them. These bipolar trans carotenoid salts
(BTCS) compounds are useful in improving diffusivity of oxygen
between red blood cells and body tissues in mammals including
humans.
BACKGROUND OF THE INVENTION
[0002] Carotenoids are a class of hydrocarbons consisting of
isoprenoid units joined in such a manner that their arrangement is
reversed at the center of the molecule. The backbone (skeleton) of
the molecule consists of conjugated carbon-carbon double and single
bonds, and can also have pendant groups. Although it was once
thought that the skeleton of a carotenoid contained 40 carbons, it
has been long recognized that carotenoids can also have carbon
skeletons containing fewer than 40 carbon atoms. The 4 single bonds
that surround a carbon-carbon double bond all lie in the same
plane. If the pendant groups are on the same side of the
carbon-carbon double bond, the groups are designated as cis; if
they are on opposite side of the carbon-carbon bond, they are
designated as trans. Because of the large number of double bonds,
there are extensive possibilities for geometrical (cis/trans)
isomerism of carotenoids, and isomerization occurs readily in
solution. A recent series of books is an excellent reference to
many of the properties, etc. of carotenoids ("Carotenoids", edited
by G. Britton, S. Liaaen-Jensen and H. Pfander, Birkhauser Verlag,
Basel, 1995 hereby incorporated by reference in its entirety).
[0003] Many carotenoids are nonpolar and, thus, are insoluble in
water. These compounds are extremely hydrophobic which makes their
formulation for biological uses difficult because, in order to
solubilize them, one must use an organic solvent rather than an
aqueous solvent. Other carotenoids are monopolar, and have
characteristics of surfactants (a hydrophobic portion and a
hydrophilic polar group). As such, these compounds are attracted to
the surface of an aqueous solution rather than dissolving in the
bulk liquid. A few natural bipolar carotenoid compounds exist, and
these compounds contain a central hydrophobic portion as well as
two polar groups, one on each end of the molecule. It has been
reported ("Carotenoids", Vol. 1A, p. 283) that carotenoid sulphates
have "significant solubility in water of up to 0.4 mg/ml". Other
carotenoids that might be thought of as bipolar are also not very
soluble in water. These include dialdehydes and diketones. A
di-pyridine salt of crocetin has also been reported, but its
solubility in water is less than 1 mg/ml at room temperature. Other
examples of bipolar carotenoids are crocetin and crocin (both found
in the spice saffron). However, crocetin is only sparingly soluble
in water. In fact, of all of the bipolar carotenoids, only crocin
displays significant solubility in water.
[0004] U.S. Pat. Nos. 4,176,179; 4,070,460; 4,046,880; 4,038,144;
4,009,270; 3,975,519; 3,965,261; 3,853,933; and 3,788,468 relate to
various uses of crocetin.
[0005] U.S. Pat. No. 5,107,030 relates to a method of making
2,7-dimethyl-2,4,6-octatrienedial and derivatives thereof.
[0006] U.S. Pat. No. 6,060,511 relates to trans sodium crocetinate
(TSC) and its uses. The TSC is made by reacting naturally occurring
saffron with sodium hydroxide followed by extractions.
[0007] In Roy et al, Shock 10, 213-7. (1998), hemorrhaged rats (55%
blood volume) were given a bolus of trans sodium crocetinate (TSC)
after 10 minutes, followed by saline after another 30 minutes. All
of the TSC-treated animals lived, while all controls died.
Whole-body oxygen consumption increased in the TSC group, reaching
75% of normal resting value after about 15 minutes.
[0008] Laidig et al, J Am Chem. Soc. 120, 9394-9395 (1998), relates
to computational modeling of TSC. A simulated TSC molecule was
"hydrated" by surrounding it with water molecules. The hydrophobic
ordering of the water in the vicinity of the TSC made it easier for
oxygen molecules to diffuse through the system. The computational
increase in diffusivity of 30% was consistent with results obtained
in both in vitro and animal experiments.
[0009] In Singer et al, Crit Care Med 28, 1968-72. (2000), TSC
improved hemodynamic status and prolonged rat survival in a rat
model of acute hypoxia. Hypoxia was induced using a low oxygen
concentration (10%) air mixture: after 10 minutes the animals were
given either saline or TSC. Hypoxemia led to a reduction in blood
flow, and an increase in base deficit. Only 2 of 6 animals survived
in the control group. The treated group all survived with good
hemodynamic stability for over two hours, with a slow decline
thereafter.
SUMMARY OF THE INVENTION
[0010] The subject invention relates to bipolar trans carotenoid
salts (BTCS) compounds and synthesis of such compounds having the
structure:
YZ-TCRO-ZY
[0011] where:
[0012] Y=a cation
[0013] Z=polar group which is associated with the cation, and
[0014] TCRO=trans carotenoid skeleton.
[0015] The subject invention also relates to individual BTCS
compound compositions (including a TSC composition) wherein
absorbency of the highest peak (of an aqueous solution of the BTCS
composition) which occurs in the visible wave length range divided
by the absorbency of the peak which occurs in the UV wave length
range, is greater than 8.5, advantageously greater than 9, most
advantageously greater than 9.5.
[0016] The invention also relates to a method of treating a variety
of diseases comprising administering to a mammal in need of
treatment a therapeutically effective amount of a compound having
the formula:
YZ-TCRO-ZY
[0017] The invention also includes several methods of solubilizing
and synthesizing compounds having the formula:
YZ-TCRO-ZY
[0018] The invention also relates to an inhaler for delivery of the
compounds of the invention.
DETAILED DESCRIPTION OF THE INVENTION
[0019] A new class of carotenoid and carotenoid related compounds
has been discovered. These compounds are referred to as "bipolar
trans carotenoid salts" (BTCS).
COMPOUNDS OF THE INVENTION
[0020] The subject invention relates to a class of compounds,
bipolar trans carotenoid salts, that permit the hydrophobic
carotenoid or carotenoid related skeleton to dissolve in an aqueous
solution, and methods for making them. The cations of these salts
can be a number of species, but advantageously sodium or potassium
(these are found in most biological systems). Commonly owned U.S.
Pat. No. 6,060,511, hereby incorporated by reference in its
entirety, describes an extraction method for making trans sodium
crocetinate, TSC (a BTCS) starting from saffron.
[0021] A general structure for a bipolar trans carotenoid salt
is:
YZ-TCRO-ZY
[0022] where:
[0023] Y (which can be the same or different at the two ends)=a
cation, preferably Na.sup.+ or K.sup.+ or Li.sup.+. Y is
advantageously a monovalent metal ion. Y can also be an organic
cation, e.g., R.sub.4N.sup.+, R.sub.3S.sup.+, where R is H, or
C.sub.nH.sub.2n+1 where n is 1-10, advantageously 1-6. For example,
R can be methyl, ethyl, propyl or butyl.
[0024] Z (which can be the same or different at the two ends)=polar
group which is associated with the cation. Optionally including the
terminal carbon on the carotenoid (or carotenoid related compound),
this group can be a carboxyl (COO.sup.-) group or a CO group. This
group can also be a sulfate group (OSO.sub.3.sup.-) or a
monophosphate group (OPO.sub.3.sup.-), (OP(OH)O.sub.2.sup.-), a
diphosphate group, triphosphate or combinations thereof.
[0025] TCRO=trans carotenoid or carotenoid related skeleton
(advantageously less than 100 carbons) which is linear, has pendant
groups (defined below), and typically comprises "conjugated" or
alternating carbon-carbon double and single bonds (in one
embodiment, the TCRO is not fully conjugated as in a lycopene). The
pendant groups are typically methyl groups but can be other groups
as discussed below. In an advantageous embodiment, the units of the
skeleton are joined in such a manner that their arrangement is
reversed at the center of the molecule. The 4 single bonds that
surround a carbon-carbon double bond all lie in the same plane. If
the pendant groups are on the same side of the carbon-carbon double
bond, the groups are designated as cis; if they are on the opposite
side of the carbon-carbon bond, they are designated as trans. The
compounds of the subject invention are trans. The cis isomer
typically is a detriment--and results in the diffusivity not being
increased. In one embodiment, a trans isomer can be utilized where
the skeleton remains linear.
[0026] Examples of trans carotenoid or carotenoid related skeletons
are: 1
[0027] where pendant groups X (which can be the same or different)
are hydrogen (H) atoms, or a linear or branched group having 10 or
less carbons, advantageously 4 or less, (optionally containing a
halogen), or a halogen. Examples of X are a methyl group
(CH.sub.3), an ethyl group (C.sub.2H.sub.5), a halogen-containing
alkyl group (C.sub.1-C.sub.10) such as CH.sub.2Cl, or a halogen
such as Cl or Br. The pendant X groups can be the same or different
but the X groups utilized must maintain the skeleton as linear.
[0028] Although many carotenoids exist in nature, carotenoid salts
do not. Commonly owned U.S. Pat. No. 6,060,511 relates to trans
sodium crocetinate (TSC). The TSC was made by reacting naturally
occurring saffron with sodium hydroxide followed by extractions
that selected primarily for the trans isomer.
[0029] The presence of the cis and trans isomers of BTCS can be
determined by looking at the ultraviolet-visible spectrum for the
carotenoid sample dissolved in an aqueous solution. Given the
spectrum, the value of the absorbency of the highest peak which
occurs in the visible wave length range of 416 to 423 nm (the
number depending on the solvent used) is divided by the absorbency
of the peak which occurs in the UV wave length range of 250 to 256
nm, can be used to determine the purity level of the trans isomer.
When the BTCS is dissolved in water, the highest visible wave
length range peak will be at about 421 nm and the UV wave length
range peak will be at about 254 nm. According to M. Craw and C.
Lambert, Photochemistry and Photobiology, Vol. 38 (2), 241-243
(1983) hereby incorporated by reference in its entirety, the result
of the calculation (in that case crocetin was analysed) was 3.1,
which increased to 6.6 after purification.
[0030] Performing the Craw and Lambert analysis on the trans sodium
crocetin of commonly owned U.S. Pat. No. 6,060,511 (TSC made by
reacting naturally occurring saffron with sodium hydroxide followed
by extractions which selected primarily for the trans isomer), the
value obtained is typically around 7-7.5 (a value of 8.4 was
observed once). Performing that test on the synthetic TSC of the
subject invention, that ratio is typically greater than 8.5 (e.g.
8.5 to 10), advantageously greater than 9 (e.g. 9-10), most
advantageously greater than 9.5. For the TSC synthesized according
to the improved method of Example 5, the ratio is typically greater
than 9.5 (e.g. 9.5-12). The synthesized material is a "purer" or
highly purified trans isomer.
[0031] It has been found, recently, that TSC has an aqueous
solubility of greater than 10 mg/ml at room temperature, which is
remarkable for a molecule containing such a long, hydrophobic
portion. TSC has also been found to increase the diffusivity of
oxygen through liquids.
[0032] U.S. Pat. No. 6,060,511 describes an extraction method for
making TSC starting from saffron; however, other bipolar carotenoid
salts cannot be made using that same procedure since the use of
saffron allows only a single carotenoid skeleton to be incorporated
into the salt.
[0033] The invention disclosed herein allows the synthesis of a
whole class of compounds: bipolar trans carotenoid salts which
contain various carotenoid or carotenoid related skeletons. Such
compounds are soluble in aqueous solutions and have advantageous
biological uses, such as causing an increase in oxygen utilization.
It is believed that this increase is a result of the ability of the
hydrophobic portion (the skeleton) of the bipolar trans carotenoid
salt to affect the bonding of nearby water molecules. This, in
effect, allows the oxygen molecule to diffuse faster while in this
vicinity.
SOLUBILIZING THE COMPOUNDS AND COMPOSITIONS OF THE INVENTION
[0034] The invention allows for the dissolution of a trans
carotenoid or carotenoid related skeleton molecule in aqueous
solutions. The novel methods of dissolution are related below. The
methods apply to any bipolar trans carotenoid salt and composition
thereof.
[0035] BTCS-Containing Saline Infusion Solutions
[0036] Large volumes (as much as 3 times the estimated blood loss)
of isotonic saline (also called normal saline) are infused as a
treatment for hemorrhagic shock. The isotonic saline contains 9 g
NaCl per liter of water so as not to disturb the ionic strength of
the plasma once it is infused into the body. Adding TSC to the
saline has been shown to result in a superior infusion fluid,
however, one cannot simply mix TSC powder with the saline to make
such a solution. About 50% of the TSC dissolves in normal saline no
matter how much TSC is added (up to several milligrams per ml),
which means that undissolved particles of TSC are still present. In
order to prevent that, a stock solution can be made by adding more
than twice the amount of TSC needed and then centrifuging out the
particles that do not dissolve. The actual composition of the stock
solution can be verified using UV-visible spectroscopy. This stock
solution can be added to normal saline and the TSC remains
dissolved.
[0037] This method can be used to dissolve a BTCS in other types of
sodium chloride solutions, as well as in solutions of other salts
such as KCl, Na.sub.2SO.sub.4, lactate, etc. Several, eg 1-3 mg/ml,
can be put into solution in this manner.
[0038] Dilute Solution of Sodium Carbonate Dissolves BTCS
[0039] A BTCS such as TSC dissolves in very dilute sodium carbonate
solutions. A dilute, eg 0.00001-0.001M, solution of sodium
carbonate can be added, dropwise, to deionized water until the pH
is 8.0 (the pH of deionized water is usually 5-6). This only takes
a few drops of the very dilute sodium carbonate per, say, 50 mls of
deionized water. This sodium carbonate-deionized water solution is
capable of completely dissolving a large amount of TSC (greater
than 10 mg/ml)--which is remarkable considering the hydrophobicity
of the carotenoid portion of the BTCS.
[0040] A BTCS can be supplied as a powder along with a sterilized
bottle of the sodium carbonate water. This concentrated solution
can then be injected directly (very small volumes of solutions
having a lower ionic strength than plasma can be injected), or the
concentrated solution can be added to normal saline and then
injected. If TSC is dissolved in the sodium carbonate-water solvent
and then more of the same solvent is added--the TSC stays in
solution.
[0041] In another embodiment, sodium bicarbonate is used instead of
sodium carbonate. Other salts which result in the deionized water
having a basic pH can also be used.
[0042] Carotenoid skeleton concentrations of 5-10 mg/ml can be
achieved with this procedure.
[0043] Water Dissolves BTCS
[0044] Although TSC dissolves in water (tap, distilled, deionized),
these solutions are only stable if the pH is adjusted so as to make
the solution basic. TSC is more soluble in deionized water (very
few Na.sup.+ ions present) than in normal water. A BTCS, such as
TSC, will dissolve in just deionized water alone, but, if plain
deionized water is added to that solution, the TSC will precipitate
out. A BTCS will dissolve in just deionized water alone, but
additional deionized water may cause precipitation of the BTCS if
the pH is not adjusted to make it slightly basic.
[0045] Other Methods of Solubilizing BTSC
[0046] The BTCS can be formulated in a delivery system that
enhances delivery. See Formulations of the Compounds of the
Invention below.
SYNTHESIS OF THE COMPOUNDS OF THE INVENTION
[0047] Bipolar Trans Carotenoid Salts
[0048] Set forth below are the novel synthesis methods that can be
used for synthesizing bipolar trans carotenoid salts. There can be
variations in various steps of the synthesis that are obvious to
one skilled in the art.
[0049] A. TSC Synthesis
[0050] Trans sodium crocetinate (TSC) can be synthesized by
coupling a symmetrical C.sub.10 dialdehyde containing conjugated
carbon-carbon double bonds (2,7-dimethylocta-2,4,6-triene-1,8-dial)
with [3-carbomethoxy-2-buten-1-ylidene] triphenylphosphorane. This
results in the formation of a trans dimethyl ester of crocetin.
This dimethyl ester is then converted to the final TSC product by
saponification. Typically, saponification is accomplished by
treating an ester with either aqueous sodium hydroxide or sodium
hydroxide dissolved in THF (tetrahydrofuran); however, these
methods did not give the best results in this case. Saponification
can be accomplished very well, in this case, by reacting the ester
with an NaOH/methanol solution. After saponification, the TSC is
recovered by drying in a vacuum.
[0051] The C.sub.10 dialdehyde and the triphenylphospborane
reactants used in this synthesis can be made via different routes.
For example, the C.sub.10dialdehyde was prepared starting with
ethyl bromoacetate and furan using Wittig chemistry. Tiglic acid
was the starting material for making the desired phosphorane.
Different lengths of carotenoid skeletons can be made by joining
together reactants of different lengths (for example a C.sub.14
dialdehyde and triphenylphosphorane). This procedure results in the
formation of different trans bipolar carotenoid salts. Alterations
can also be made so as to obtain different pendant groups (TSC has
methyl groups for the pendant groups).
[0052] The TSC made in this manner is soluble in water (pH adjusted
to 8.0 with a very dilute solution of sodium carbonate) at a level
>10 mg/ml at room temperature. Other bipolar trans carotenoid
salts are soluble at room temperature in water having a pH that is
neutral or above. As used herein, "soluble" means that amounts
greater than 5 mg will dissolve per ml of water at room temperature
(as noted previously, carotenoid references state that 0.4 mg/ml is
"highly significant solubility"--but that is lower than the subject
definition of solubility).
[0053] B. General Synthesis
[0054] Carotenoid or carotenoid related structures can be built up
in the following manner: 2
[0055] (3-carbomethoxy-2-buten-1-ylidene)triphenylphosphorane (or a
related compound when X is other than a methyl group) is a key
precursor to add isoprenoid units (or isoprenoid related units) to
both ends of a symmetrical carotenoid (or carotenoid related
compound). This process can be repeated infinitely. For example,
dimethyl trans crocetinate can be reduced to the corresponding
symmetrically dialdehyde using the chemistry described above. This
dialdehyde can be reacted with excess
(3-carbomethoxy-2-buten-1-ylidene)triphenylphosphorane to give the
corresponding diester. This synthetic sequence can be repeated
again and again.
[0056] Improved Synthesis
[0057] 2,7-Dimethyl-2,4,6-octatrienedial is a key intermediate
toward the synthesis of TSC. This key precursor has three double
bonds and thus several isomers are possible. For TSC, the all trans
isomer (E,E,E-isomer) is required. The general synthesis route
involves an 11-step synthesis with relatively low yields and poor
selectivity in several steps (see Example 1). As a result, column
chromatography is required to purify several intermediates along
the way.
[0058] The improved synthesis route is much simpler (see the
reaction scheme below). The 3-step process as described in U.S.
Pat. No. 5,107,030, hereby incorporated by reference in its
entirety, gives a mixture of geometric isomers of the dialdehyde
(U.S. Pat. No. 5,107,030 does not note this mixture). In the method
of the subject invention described in Example 1, 96-97% of the
desired isomer (all trans or E,E,E-isomer) is obtained by several
recrystallizations from methanol or ethyl acetate in 59% yield.
[0059] The improved synthesis method of the subject invention
involves converting the remaining isomeric mixture of dialdehydes
into the desired trans aldehyde (E,E,E) by isomerization with a
sulfinic acid (RSO2H where R is C1 through C.sub.10 straight or
branched alkyl group or an aryl group (a substituted phenyl group)
such as para-toluenesulfinic acid, in an appropriate solvent such
as 1,4-dioxane, tetrahydrofuran or dialkyl ether where the alkyl
group is one or two of a C1 through C.sub.10 straight or branched
alkyl group. An additional 8% yield of the pure desired dialdehyde
is obtained, raising the overall yield of the last step from 59% to
67% yield. This yield improvement is important. This isomerization
step can be incorporated into the third step of the method of U.S.
Pat. No. 5,107,030 to get a better yield.
[0060] Improved Synthesis Route: 3
[0061] Two Undesired Isomers: 4
[0062] Isomerization of Undesired to Desired Dialdehdye: 5
[0063] Saponification can be accomplished by dissolving the diester
in methanol and then adding a base such as NaOH (Y of the BTCS is
then Na.sup.+). Alternatively, the diester can be dissolved in
methanol already containing the base. The NaOH is typically aqueous
(20-60% by wt.) but can be solid. Alternatives to methanol for
dissolving the diester are ethanol, propanol and isopropanol.
Saponification can be carried out in various ways commercially. A
one or two phase system (one organic and one aqueous phase) can be
used.
[0064] Trans crocetin can also be synthesized according to the
methods described above.
[0065] In addition, as has been reported for TSC, such BTCS
compounds increase the diffusivity of oxygen through water (this
will also depend on the nature of the hydrophobic portion
incorporated into the final product such as carbon chain length)
since it is believed that the hydrophobic interactions of the
carotenoid skeleton with water result in the increased
diffusivity).
FORMULATIONS OF THE COMPOUNDS OF THE INVENTION
[0066] A concentrated solution of a bipolar trans carotenoid salt
can be made, as described previously, by dissolving it in a very
dilute solution of sodium carbonate. The resulting mixture can then
be used in that manner, or can be diluted further with normal
saline or other aqueous solvents. In addition, solutions of a
bipolar trans carotenoid salt can be made by dissolving the bipolar
trans carotenoid salt directly in a salt solution and then getting
rid of any material that does not dissolve.
[0067] The bipolar trans carotenoid salts are stable in a dry form
at room temperature, and can be stored for long periods.
Advantageously, a formulation of such salts, if given orally, is
absorbed in the gut, rather than the stomach.
[0068] Although the compounds of the invention can be administered
alone, they can be administered as part of a pharmaceutical
formulation. Such formulations can include pharmaceutically
acceptable carriers known to those skilled in the art as well as
other therapeutic agents-see below. Advantageously, the formulation
does not include a compound that inhibits the ability of the
compounds of the invention to improve diffusivity of oxygen.
[0069] Appropriate dosages of the compounds and compositions of the
invention will depend on the severity of the condition being
treated. For a dose to be "therapeutically effective", it must have
the desired effect, i.e. increase the diffusivity of oxygen. This
in turn, will cause oxygen-related parameters to return towards
normal values.
[0070] Administration can be by any suitable route including oral,
nasal, topical, parenteral (including subcutaneous, intramuscular,
intravenous, intradermal and intraosseus), vaginal or rectal. The
preferred route of administration will depend on the circumstances.
An inhalation route is advantageous for treatment in emergency
situations, where it is necessary for the BTCS to enter the
bloodstream very quickly. The formulations thus include those
suitable for administration through such routes (liquid or powder
to be nebulized). It will be appreciated that the preferred route
may vary, for example, with the condition and age of the patient.
The formulations can conveniently be presented in unit dosage form,
e.g., tablets and sustained release capsules, and can be prepared
and administered by methods known in the art of pharmacy. The
formulation can be for immediate, or slow or controlled release of
the BTCS. See for example, the controlled release formulation of WO
99/15150 hereby incorporated by reference its entirety.
[0071] Formulations of the present invention suitable for oral
administration can be presented as discrete units such as pills,
capsules, cachets or tablets, as powder or granules, or as a
solution, suspension or emulsion. Formulations suitable for oral
administration further include lozenges, pastilles, and inhalation
mists administered in a suitable base or liquid carrier.
Formulations for topical administration to the skin can be
presented as ointments, creams, gels, and pastes comprising the
active agent and a pharmaceutically acceptable carrier or in a
transdermal patch.
[0072] Formulations suitable for nasal administration wherein the
carrier is a solid include powders of a particular size that can be
administered by rapid inhalation through the nasal passage.
Suitable formulations wherein the carrier is a liquid can be
administered, for example as a nasal spray or drops.
[0073] Formulations suitable for parenteral administration include
aqueous and non-aqueous sterile injection solutions that can
contain antioxidants, buffers, bacteriostats and solutes which
render the formulation isotonic with the blood of the intended
recipient, and aqueous and nonaqueous sterile suspensions which can
include suspending agents and thickening agents. The formulations
can be presented in unit or multi-dose containers, for example
sealed ampules and vials, and can be lyophilized, requiring only
the addition of the sterile liquid carrier such as water for
injection immediately prior to use. Injection solutions and
suspensions can be prepared from sterile powders, granules and
tablets.
USES OF THE COMPOUNDS AND COMPOSITIONS OF THE INVENTION
[0074] A wide variety of conditions are controlled or are mediated
by delivery of oxygen to body tissues. The compounds and
compositions of the subject invention can be used in the same
pharmaceutical applications described for crocetin in the same
effective amounts; see U.S. Pat. Nos. 4,176,179; 4,070,460;
4,046,880; 4,038,144; 4,009,270; 3,975,519; 3,965,261; 3,853,933;
and 3,788,468 each of which is hereby incorporated by reference in
its entirety.
[0075] TSC has been shown to increase the diffusivity of oxygen
through aqueous solutions by about 30%. Thus, the compounds of the
invention are useful for treating diseases/conditions which are
characterized by low oxygen (hypoxia) such as respiratory diseases,
hemorrhagic shock and cardiovascular diseases, atherosclerosis,
emphysema, asthma, hypertension, cerebral edema, papillomas, spinal
cord injuries, among others. Other bipolar trans carotenoid salts
have similar properties. Such compounds can also be used in
conjunction with other methods commonly suggested for increasing
oxygen utilization in the body, such as oxygen therapy and the use
of hemoglobins or fluorocarbons.
[0076] In one embodiment of the invention, a BTCS is administered
to the patient while administering oxygen. Alternatively,
hemoglobins or fluorocarbons and a BTSC can be given together. In
these cases, an additive effect is realized.
[0077] The minimum dosage needed for treatment for any of these
salts is that at which the diffusivity of oxygen increases. The
effective dosage of the compounds of the inventions will depend
upon the condition treated, the severity of the condition, the
stage and individual characteristics of each mammalian patient
addressed. Dosage will vary, however, from about 0.001 mg of active
compound per kg of body weight up to about 500 mg per kg, and
advantageously from about 0.01-30 mg/kg of body weight. IV
administration is advantageous but other routes of injection can
also be used such as intramuscular, subcutaneous or via inhalation.
Oral administration can also be used as can transdermal delivery or
intraosseus delivery.
[0078] Respiratory Disorders
[0079] Bipolar trans carotenoid salts can be used to treat
respiratory disorders. These are described as conditions in which
the arterial partial pressure of oxygen is reduced, such as value
of 60 to 70 mm Hg rather than the normal value of 90-100 mm Hg.
Such diseases are emphysema, acute respiratory distress syndrome
(ARDS) or chronic obstructive pulmonary disease (COPD).
[0080] TSC increases the value of the partial pressure of oxygen in
the blood when it is low (this is a symptom of emphysema, ARDS and
COPD). Increasing the partial pressure of oxygen in the blood
relieves many of the symptoms of emphysema, ARDS and COPD. TSC does
not cure the cause of the disease, but relieves the oxidative
distress and damage resulting from that underlying cause.
[0081] Hemorrhagic Shock
[0082] Hemorrhagic shock is marked by a decrease in oxygen
consumption. Bipolar trans carotenoid salts increase the body's
oxygen consumption by causing more oxygen to diffuse from the red
blood cells to the tissues. TSC has been shown to increase the
oxygen consumption of rats undergoing hemorrhagic shock, and has
also been shown to offset other symptoms of shock. The compounds of
the invention cause the low blood pressure to increase, reduce the
increased heart rate, and reverse the blood acidosis that develops
during shock. The compounds of the invention also reduce organ
damage subsequent to hemorrhagic shock.
[0083] The compounds of the invention can be used for hemorrhagic
shock by administering them by inhalation, injecting them, or by
adding them to a standard resuscitation fluid (Ringer's lactate or
normal saline).
[0084] Cardiovascular Disease
[0085] In western culture, the leading cause of death is ischemic
heart disease. Death may result from either a gradual deterioration
of the ability of the heart to contract or, frequently, a sudden
stoppage. Sudden cardiac death (SCD) covers the time period
beginning 60 seconds after symptoms begin to 24 hours later. These
deaths are usually a consequence of acute coronary occlusion
(blockage) or of ventricular fibrillation (which can result from
the occlusion).
[0086] Myocardial ischemia exists when there is an insufficient
supply of oxygen to the cardiac muscle. When coronary blood flow is
extremely low, cardiac muscle cannot function and dies. That area
of muscle is said to be infarcted. Most often, diminished coronary
blood flow is caused by atherosclerosis that occurs in the coronary
arteries. Ischemia results in impaired mechanical and electrical
performance and muscle cell injury, which may lead to a lethal
arrhythmia, called ventricular fibrillation (VF). In ventricular
fibrillation, the electrical activity of the ventricles of the
heart is chaotic and results in an electrocardiogram with an
erratic rhythm and no recognizable patterns. Ventricular
fibrillation occurs frequently with myocardial ischemia and
infarction and is nearly always the cause of sudden cardiac death.
Bipolar trans carotenoid salts are beneficial in treating
myocardial ischemia. Atherosclerosis, which is frequently a
precursor to myocardial infarction, can also be treated with these
salts.
[0087] Ischemia
[0088] Bipolar trans carotenoid salts are also beneficial in
treating other forms of ischemia (insufficient blood flow to
tissues or organs) such as kidney, liver, spinal cord, and brain
ischemia including stroke.
[0089] Hypertension
[0090] Hypertension, or high blood pressure, is frequently
associated with cardiovascular disease. The compounds of the
invention can be used to reduce blood pressure.
[0091] Performance Enhancement
[0092] BTCS enhance aerobic metabolism, increasing oxygen
consumption levels during walking, running, lifting, etc. Endurance
is also increased.
[0093] Traumatic Brain Injury
[0094] Hypoxia following traumatic brain injury results in
increased brain damage. BTCS increase oxygen levels in brain tissue
after impact injury (focal or diffuse injury). Examples of impact
injury include car/motorcycle accidents and falls. BTCS also
augment the amount of oxygen reaching normal brain tissue when
hyper-oxygen therapy is used.
[0095] Alzheimer's Disease
[0096] BTCS increase brain oxygen consumption levels in Alzheimer's
Disease, thus alleviating symptoms of Alzheimer's Disease. Blood
flow and oxygen consumption decline to level some 30% below that
seen in non-demented elderly people Wurtman, Scientific American,
Volume 252, 1985.
[0097] The increased oxygen consumption levels in the brain created
by BTCS also reduce memory loss.
[0098] Diabetes
[0099]
[0100] BTCS are useful for treating complications of diabetes such
as ulcers, gangrene and diabetic retinopathy. Diabetic foot ulcers
heal better with hyperbaric oxygen breathing treatment, M. Kalani
et al. Journal of Diabetes & Its Complications, Vol 16, No. 2,
153-158, 2002.
[0101] BTCS also help the complication of diabetic retinopathy
which is related to low oxygen tension, Denninghoff et al.,
Diabetes Technology & Therapeutics, Vol. 2, No. 1, 111-113,
2000.
[0102] Other Uses
[0103] Bipolar trans carotenoid salts can also be used for the
treatment of spinal cord injury, cerebral edema, and skin
papillomas. In all cases, they alleviate the condition, making it
less severe. It is believed that this results from the increase in
oxygen consumption that results from the use of bipolar trans
carotenoid salts.
[0104] BTCS also scavenge oxygen-derived free radicals.
[0105] The following Examples are illustrative, but not limiting of
the compositions and methods of the present invention. Other
suitable modifications and adaptations of a variety of conditions
and parameters normally encountered which are obvious to those
skilled in the art are within the spirit and scope of this
invention.
EXAMPLES
Example 1
[0106] Synthesis of Trans Sodium Crocetinate 6
[0107] Trans sodium crocetinate is synthesized by coupling a
symmetrical C.sub.10 dialdehyde containing conjugated carbon-carbon
double bonds with
[3-carbomethoxy-2-buten-1-ylidene]triphenylphosphorane. This
product is then saponified using a solution of NaOH/methanol.
[0108] To ethyl bromoacetate, trephenylphosphine dissolved in ethyl
acetate (at a concentration of around 2 moles/liter) is slowly
added. After isolation, and treatment with base, the product can be
treated with methyl iodide, followed by caustic, to form the
phosphorane. The basic compound to form the carotenoid skeleton can
be made starting with a ring compound such as furan in this case.
Furan is reacted with bromine and methanol, followed by a selective
deprotonation step to form a monoaldehyde. This is then coupled
with the phosphorane. Acidic conditions deprotected the other
dimethyl acetal group to afford the free aldehyde. This compound is
then reacted again with the same phosphorane to give the diethyl
diester. The ester groups are reduced to alcohols, and subsequent
oxidation (such as with MnO.sub.2) results in the C.sub.10 skeleton
in the dialdehyde form. This is later reacted with a phosphorane
made from tiglic acid. The tiglic acid is esterified with methanol
under acidic conditions to give the methyl ester, followed by a
bromination step. The resulting allylic bromide isomers are formed,
and can be separated using crystallization. Subsequent treatment of
the desired bromide with sodium hydroxide results in the desired
phosphorane. This phosphorane and the C.sub.10 dialdehyde are then
dissolved in a solvent such as toluene or benzene and refluxed. The
resulting product isolated as a powder and is then saponified with
a 40% NaOH/methanol mixture to form the TSC after solvent
removal.
[0109] Trans-sodium crocetinate 1 (TSC) was prepared in a 17 step
synthetic sequence in an overall yield of 1.5%. A total of 4.1 g of
TSC was prepared with ethyl bromoacetate, furan and tiglic acid as
starting materials. 7
[0110] Trans-sodium crocetinate (TSC) was synthesized from
saponification of dimethyl crocetinate, the preparation of which
was based on a total synthesis reported by Buchta and Andree..sup.1
The synthetic strategy behind preparing dimethyl crocetinate was
based on coupling the symmetrical C.sub.10 dialdehyde
(2,7-dimethylocta-2,4,6-triene-1,8-dial) with
(3-carbomethoxy-2-buten-1-ylidene)triphenylphosphorane. 8
[0111] Although the original Buchta and Andree article.sup.1 was
titled "The Total Synthesis of
trans-2,2-Bisdimethyl-crocetin-dimethyl ester and
trans-Crocetin-dimethyl ester," experimental details and yields
were not reported. Procedures for the various steps leading to the
C.sub.10 dialdehyde and phosphorane were found after an extensive
survey of the literature. Ultimately, TSC was prepared in a 17 step
sequence with ethyl bromoacetate, furan and tiglic acid as the
starting materials in an overall yield of 1.5%.
[0112] The C.sub.10 symmetrical dialdehyde was prepared from ethyl
bromoacetate.sup.2 and furan.sup.3 using Wittig chemistry. Ethyl
bromoacetate was treated with triphenylphosphine and methyl iodide
to give the phosphorane 6: 9
[0113] The yield for the first step was a respectable 92%.
Quantitation of the subsequent steps of this sequence were
complicated by the nature of phosphorane 4 and phosphonium salt 5.
Both of these compounds were extremely viscous syrups which foamed
vigorously while concentrating on a rotary evaporator. Both
compounds could be conveniently handled as methylene chloride
solutions and the overall yield of phosphorane 6 appeared to be
acceptable from a qualitative point of view (estimated at better
than 75%).
[0114] Furan was ring-opened with bromine to afford fumaraldehyde
bis(dimethylacetal) 8..sup.3 10
[0115] Mono-deprotection of bis(dimethylacetal) 8 under acidic
conditions.sup.4 gave aldehyde 9, which was then coupled with
phosphorane 6 to give 10 in a 45% yield. Acidic conditions were
used to deprotect the dimethylacetal 10. Treating 11 with
phosphorane 6 gave diester 12. The ester groups were reduced to
alcohols by DIBAL-H and subsequent oxidation with MnO.sub.2 gave
the C.sub.10 dialdehyde 14. The trans stereochemistry of 14 was
determined by NMR data. In particular, the C.sub.2 symmetry of the
compound gave the expected 5 resonances in the .sup.13C NMR
spectrum and the .sup.1H NMR spectrum showed signals at .delta.
9.54 (1H), 7.07 (2H) and 1.95 (3H). 11
[0116] The range in yields of steps h-k reflect improvements in
isolation from intial pilot studies to scaled up reactions.
[0117] Tiglic acid 15 was converted to phosphorane 20 in a 4 step
sequence. Fisher esterification conditions on 15 gave the methyl
ester 16. Reaction with NBS gave a mixture of 59% methyl
.gamma.-bromotiglate, 26% methyl .alpha.-bromotiglate and the
balance of the material was unreacted starting material. The
formation of regioisomers was expected based on the reported
literature..sup.5 In the following step, the .alpha./.gamma.
mixture of phosphonium salts was recrystallized to give the desired
.gamma.-phosphonium bromide 19..sup.6 Subsequent treatment with
sodium hydroxide gave the phosphorane 20. 12
[0118] Phosphorane 20 and C.sub.10 dialdehyde 14 were coupled by
refluxing in benzene..sup.6 Dimethyl crocetinate 21 was isolated as
a red powder. Saponification of the methyl ester proved to be more
difficult than expected. Treating the ester 21 with 2 eq. NaOH in
THF/H.sub.2O at r.t. and reflux left the material unchanged.
Solubility appeared to be a significant problem, so pyridine was
added. While this did dissolve most of the solids, refluxing a
mixture of pyridine and 2.5 N NaOH yielded no product. Standard
THF/2.5 N NaOH saponification conditions also had no effect on the
ester. Eventually, 40% NaOH/methanol at reflux for an overnight
period proved to be successful. This gave TSC 1 as an orange solid.
13
[0119] Attempts were made to dissolve TSC in order to obtain a
.sup.1H NMR spectrum. However, TSC was practically insoluble in
most common organic solvents (chloroform, DMSO, pyridine, methanol,
acetone, and glacial acetic acid). The TSC produced from this
project was characterized by IR, UV, HPLC and elemental analyses.
IR showed characteristic absorbance at 1544 and 1402 cm.sup.-1
(consistent with conjugated carboxylates). UV and HPLC were
consistent with authentic TSC..sup.7 Elemental analyses gave
satisfactory values.
[0120] The overall yield of the reaction sequence was 1.5% (based
on furan).
[0121] The synthesis is described in detail below:
[0122] All reagents and chemicals were purchased from Aldrich or
Sigma and used as received unless stated otherwise. Solvents were
purchased from Fisher Scientific as ACS reagent or HPLC grade and
used without further purification. Anhydrous solvents were
purchased from Aldrich in Sure/Seal.TM. bottles and used directly
without further purification. Deionized water was obtained from an
in-house Culligan water treatment system.
[0123] Melting points were obtained on a Mel-Temp II and were
uncorrected. Infrared spectra were measured on a Perkin-Elmer 1600
FTIR spectrophotometer. Nuclear magnetic spectra were measured on a
JEOL FX90Q spectrometer using a 5 mm multinuclei probe with
internal or external deuterium lock depending on the nature of the
sample. Proton and carbon NMR chemical shifts were assigned
relative to TMS or the deuterated solvent respectively. Phosphorus
NMR spectra were generally run in the proton-decoupled mode with a
coaxial insert tube of 5% aqueous phosphoric acid as the external
standard.
[0124] Routine analyses by gas chromatography to evaluate reaction
progress or estimate product composition were conducted on a Varian
3700 gas chromatograph equipped with a flame ionization detector
and a Hewlett Packard 3394A integrator. A 1 microliter solution was
injected onto a 15 meter DB5 column (0.53 mm ID and 1.5 micron film
thickness) with helium carrier gas using a temperature program from
50 to 250.degree. C. at 20.degree. C./min with a 10 minute hold at
250.degree. C. The injector and detector temperatures were
typically set at 250.degree. C.
[0125] Thin layer chromatography was conducted on Baker-flex
2.5.times.7.5 cm silica gel plates with or without fluorescent
indicator (1B2 or 1B2-F) depending on the method of detection. The
components on the developed plates were detected by UV.
[0126] Elemental analyses were conducted by Quantitative
Technologies, Inc., Whitehouse, N.J.
[(Ethoxycarbonyl)methylene]triphenylphosphorane (4).sub.2
(ACL-G29-1)
[0127] Triphenyl phosphine (235.6 g, 0.90 mol) was dissolved in
EtOAc (540 mL). Approximately 30 min was required for all of the
solids to dissolve. The process was endothermic (solution cooled to
13.degree. C. when the ambient temperature was 20.degree. C.). A
solution of ethyl bromoacetate (100 mL, 0.90 mol) in EtOAc (400 mL)
was added dropwise over a 1.5 h period. A white precipitate formed
during the addition. Stirred overnight (20 h) at ambient
temperature (18.degree. C.).
[0128] The solids were collected by vacuum filtration rinsing with
copious amounts of Et.sub.2O. Dried overnight in vacuo at
45.degree. C. to give 3 as a white solid 356.3 g, 92.6% yield (0.83
mol). .sup.1H NMR was consistent with literature values.
[0129] The solid was dissolved in methylene chloride (3 L) and
treated with 1 M NaOH (3.6 L) in a 12 L flask with vigorous
stirring for 45 min. The organic layer was separated and the
aqueous phase was extracted with additional methylene chloride
(2.times.1 L). Organic layers were dried (MgSO.sub.4) and
concentrated until approximately 1 L of volume remained. A small
amount of material was removed and examined by .sup.1H NMR and
found to be consistent with literature values.
[1-(Ethoxycarbonyl)ethylidene]triphenylphosphoniun iodide (5).sub.2
(ACL-G29-2)
[0130] The material from ACL-G29-1 was treated with iodomethane
(64.0 mL, 1.03 mol) as the reaction flask was cooled in an ice
bath. The reaction mixture was checked by TLC (silica gel, 10%
MeOH/CHCl.sub.3) when the addition was completed (1 h) and it
showed a considerable amount of starting material remained. The ice
bath was removed and the reaction mixture was checked by TLC after
1.5 h, it looked complete based on a tightening of the main band
(s.m. streaked). The reaction mixture was concentrated on a rotary
evaporator, when most of the solvent was removed, the product began
foaming and creped up the vapor duct. The phosphonium salt 5
appeared was an extremely viscous syrup which was kept as a
methylene chloride solution to facilitate handling. Because of the
nature of 5, the material was not quantitated.
[1-(Ethoxycarbonyl)ethylideneltriphenylphosphorane (6).sub.2
(ACL-G29-2A)
[0131] A portion of 5 dissolved in CH.sub.2Cl.sub.2 (350 mL) and
vigorously stirred with 1 M NaOH (500 mL) for 45 min. The organic
layer was separated and the aqueous was extracted with
CH.sub.2Cl.sub.2 (2.times.100 mL). Combined organic layers were
dried (MgSO.sub.4) and concentrated to give 6 as a yellow solid,
8.0 g. .sup.1H NMR spectrum was consistent with literature
values.
Fumaraldehyde bis(dimethylacetal) (8).sub.3 (ACL-G29-3)
[0132] A solution of furan (88.0 g, 1.29 mol) in anhydrous MeOH
(650 mL) was cooled to -45.degree. C. under N.sub.2. A solution of
bromine (68.0 mL, 1.32 mol) was added dropwise over a 2.5 h period
at a rate to maintain .ltoreq.-45.degree. C. The red solution was
allowed to warm to -10.degree. C. over a 2.5 h period and held for
an additional 2 h. The reaction mixture was a pale amber color.
Addition of 5 g Na.sub.2CO.sub.3 produced a considerable amount of
outgassing and a 4.degree. C. exotherm. The reaction mixture was
cooled with dry-ice and the remaining Na.sub.2CO.sub.3 (210 g
total) was added over a 50 min period. After holding at -10.degree.
C. overnight (11 h, the cooling bath was removed and the reaction
mixture was allowed to warm to room temperature and stirred for 20
h.
[0133] The salts were removed by vacuum filtration and the filtrate
was vacuum distilled with a vigreux column until approximately 150
mL had been removed. Additional salt had precipitated out and was
causing the distillation pot to bump violently. After filtration,
another 150 mL was distilled and more salt came out of solution.
Once again, severe bumping was a problem. The still pot was cooled,
filtered, the filtrate treated with Et.sub.2O (400 mL) and the
precipitate removed by vacuum filtration. At least 120 g of salt
was collected (early crops of salt were discarded without
quantitation). The majority of the Et.sub.2O was removed on a
rotary evaporator at 25.degree. C. with a water aspirator.
Distillation was resumed with a vigreux column, 8 was collected as
a clear, colorless liquid 175.2 g (76.9% yield), b.p. 86-92.degree.
C./9 torr (lit. 85-90 C/15 torr). .sup.1H NMR spectrum was
consistent for the desired product. GC analysis: 81.9% pure.
Fumaraldyhyde mono(dimethylacetal) (9).sub.4 (AC L-G29-4)
[0134] Fumaraldyhyde bis(dimethylacetal) 8 (5.29 g, 0.03 mol) was
dissolved in acetone (120 mL). H.sub.2O (1.80 mL) and Amberlyst 15
(1.20 g) were sequentially added. The mixture was stirred
vigorously for 5 min then filtered to removed the resin. During
this time, the solution turned from colorless to yellow. The
filtrate was concentrated on a rotary evaporator at room
temperature and the light brown residue was distilled on a
kugelrohr (37.degree. C./200 millitorr) to give 9 as a yellow
liquid, 2.80 g, 71.8% yield. A small amount of material was lost
when the still pot bumped at the beginning. .sup.1H NMR spectrum
was consistent for the desired product, GC analysis indicated 80%
purity.
(ACL-G29-7)
[0135] Fumaraldyhyde bis(dimethylacetal) .delta. 72.1 g, 0.41 mol)
was dissolved in acetone (1600 mL). H.sub.2O (25.0 mL) and
Amberlyst 15 (16.7 g, prewashed with acetone) was added. The
mixture was stirred vigorously for 5 min then filtered to removed
the acid resin. The reaction mixture had a slight yellow tint, much
fainter than the previous large scale prep. GC analysis indicated
34.5% product and 46.1% s.m. Treated with resin for another 5 min.
GC analysis indicated 59.5% product and 21,7% s.m. Treated with
resin for another 10 min (total time 20 min). GC analysis indicated
73,9% product and 2.0% s.m. The filtrate was concentrated on a
rotary evaporator at room temperature to give a brown oil, 54 g.
Vacuum distillation gave a yellow-green oil, 34.48 g. GC analysis
indicated 64.7% purity (8.22 min) with a major impurity of 17.5%
(9.00 min) and 6.9% (9.14 min). Net recovered yield 22.3 g (0.17
mol). Analysis of the forecut by GC showed extremely dirty
material.
(ACL-G29-13)
[0136] Amberlyst 15 (8.61 g) was stirred in acetone (100 mL) for 30
min and collected by filtration. The acetal 8 (35.0 g, 0.16 mol)
was dissolved in acetonitrile (620 mL) and while mechanically
stirred, acid resin and deionized H.sub.2O (10.0 mL, 0.55 mol) was
added. The course of the reaction was monitored by TLC (10:3
hexane:Et.sub.2O), after 15 min most of the starting material had
been converted. After 20 min, only a trace of the dimethyl acetal
was detected. The resin was removed by filtration and the filtrate
was concentrated on a rotary evaporator at .ltoreq.40.degree. C.
The crude product was loaded on a Biotage column (7.5.times.9.0 cm)
eluting with 15% Et.sub.2O in hexanes to give 19.8 g. 65%
yield.
6,6-Dimethyoxy-2-methylhexa-2,4-dienoate (10).sub.2 (ACL-G29-5)
[0137] The ylide 6 (7.80 g, 22 mmol) was dissolved in methylene
chloride (65 mL). A solution of fumaraldehyde mono(dimethylacetal)
9 (2.80 g, 17 mmol) was added and the solution was stirred
overnight. Solvent was removed at reduced pressure on a rotary
evaporator. .sup.1H NMR of the crude indicated desired product was
present. Upon standing, crystals grew (presumably
triphenylphosphine oxide). The solid (14.1 g after drying by vacuum
filtration) was slurried in petroleum ether and filtered. The
filtrate was concentrated to give a yellow oil with solids
precipitated out which was dissolve in methylene chloride (15 mL)
and chromatographed on a Biotage 4.times.7.5 cm column eluting with
methylene chloride to give 10 as a yellow oil 1.8 g, 50% yield.
.sup.1H NMR spectrum of the yellow oil was consistent literature
values, however, a trace of methylene chloride remained (0.75 eq)
so the material was place on the rotary evaporator for 45 min. Mass
was reduced to 1.5 g, 40.6% yield and the methylene chloride
resonance disappeared. GC analysis major peak at 12.6 min, 87.5%
(50.degree. C., 5 min hold, 20.degree. C./min to 250.degree. C.
final temperature).
(ACL-G29-6)
[0138] A solution of ylide 6 (59.2 g, 0.16 mol) in methylene
chloride (650 mL) was cooled in an ice bath and a solution of 9
(25.7 g, 0.19 mol) was added. The solution was stirred overnight
allowing the ice bath to melt. TLC (hexane:Et.sub.2O 10:3)
indicated at least 3 other compounds running very close to the
product. Examination of the aldehyde indicated by GC analysis 50.0%
purity. Solvent was removed to give a solid/oil mixture.
(ACL-G29-8)
[0139] Ylide 6 (59.2 g, 0.16 mol) and acetal 9 (0.19 mol) was
coupled in methylene chloride (1.1 L) and worked up as described
above to give a yellow-green oil, 80 g. A portion of the crude
reaction mixture (4.13 g of the original 80 g) was placed on the
kugelrohr and distilled at 50.degree. C./250 millitorr. A colorless
oil was condensed 2.28 g, .sup.1H NMR indicated it was the starting
aldehyde while the product 10 remained in the still pot, 1.85 g.
Volatile components were removed from the bulk of the crude product
by kugelrohr distillation at 50.degree. C./200 millitorr (net 35
g).
Ethyl 2-methyl-6-oxo-hexa-2,4-dienoate (11).sub.2 (ACL-G29-9)
[0140] Acetal 10 from the pilot still pot (ACL-G29-8, 1.85 g, 9
mmol) was dissolved in acetone (33 mL). Deionized H.sub.2O (0.50
mL) and Amberlyst 15 resin (0.35 g, prewashed with acetone) were
added. The mixture was stirred for 20 min. Filtered and
concentrated on a rotary evaporator to give a yellow-green oil,
1.53 g. Chromatographed on a 4.5.times.7 cm Biotage column eluting
with 15% Et.sub.2O in hexanes. This system gave incomplete
separation, but 0.32 g of the main component was isolated and
analyzed; .sup.1H NMR spectrum was consistent with literature data
and IR (1711, 1682 cm.sup.-1) was consistent with the desired
product. GC 95.6%. An additional 0.35 g was recovered, although it
was cross contaminated with less and more polar material. The
.sup.1H NMR spectrum indicated fairly clean material. GC 90.6%.
Yield: 42%.
Diethyl 2,7-dimethylocta-2,4,6-triene-1,8-dioate (12).sub.2
(ACL-G29-10)
[0141] The aldehyde 11 (0.65 g, 3.5 mmol) from G29-9 was dissolved
and magnetically stirred in methylene chloride. Ylide (1.59 g, 4.4
mmol) was added. The light yellow-green solution turned a darker
shade yellow within minutes. TLC after 10 min indicated starting
material was almost completely consumed. After stirring for 20 h,
the reaction mixture (brown solution) was filtered through a
pipette partially filled with silica gel. The filtrate was
concentrated to give a brown solid. The solid was dissolved in 5%
Et.sub.2O in hexanes with a small amount of CHCl.sub.3.
Chromatographed on a 4.times.7.5 cm Biotage column eluting with 5%
Et.sub.2O in hexanes. The main product was isolated as a white
crystalline solid, 045 g, 50% yield. .sup.1H NMR spectrum was
consistent with literature data.
(ACL-G29-14)
[0142] An additional amount of 12 was prepared as described above
to give 21.8 g, 81.6% after chromatographic purification. .sup.1H
NMR spectrum was consistent with the desired product.
2,7-Dimethylocta-2,4,6-triene-1,8-diol (13).sub.2 (ACL-G29-11)
[0143] The diester 12 (0.45 g, 1.8 mmol) was taken up in anhydrous
hexanes (15.0 mL). It appeared as though some of the material
dissolved, but the mixture was quite cloudy. More material appeared
to come out of solution when the mixture was cooled in a -78 C
bath. Neat DIBAL-H (2.50 mL) was dissolved in anhydrous hexanes
(total volume 10.0 mL) and a portion (approximately 2 mL) was
inadvertently siphoned into the reaction mixture as the diester was
cooled in a dry-ice bath. An additional amount of DIBAL-H solution
was added until a total of 5.0 mL (6.7 mmol) was added. The
CO.sub.2 bath was allowed to warm. After stirring for 2 h 50 min,
TLC indicated the diester was completely consumed. Bath temperature
was adjusted to -20.degree. C. allowing to warm to 0.degree. C.
over 20 min. Treated with H.sub.2O/silica gel (2 mL/7 g) mixture
for 30 min. Added K.sub.2CO.sub.3 and MgSO.sub.4. Filtered to
remove the solids and thoroughly rinsed with methylene chloride.
Concentrated to give a white solid, 0.14 g, 50% yield. Note: TLC
R.sub.f=0.21 (5% MeOH/CHCl.sub.3) is quite polar. Rinsing with
methylene chloride might not have been enough to recover all of the
product. .sup.1H NMR spectrum was consistent with literature
values.
(ACL-G29-15)
[0144] The diester (5.4 g, 21 mmol) was taken up in anhydrous
hexanes (175 mL, poor solubility), cooled in a -78.degree. C. bath
and treated with a solution of DIBAL-H (14.5 mL in 50 mL anhydrous
hexanes) over a 35 min period. Vigorous gas evolution was observed
during the addition. The color of the slurry went from white to
dark yellow initially, this lightened up as additional DIBAL-H was
added. Allowed to warm to -40.degree. C. over 2 h, then transferred
to a -28.degree. C. bath overnight. The reaction mixture was
treated with a homogeneous mixture of H.sub.2O/silica gel (4
mL/14.4 g) for 30 min. MgSO.sub.4 (7.5 g) and K--.sub.2CO.sub.3
(5.1 g) was added and the reaction mixture was removed from the
cooling bath. Stirred 20 min, then filtered on a sintered glass
funnel. The solids were washed with methylene chloride--this caused
a considerable amount of precipitate to form. Warming while placed
on a rotary evaporator dissolved the precipitated solids. The
solids remaining on the sintered glass funnel was washed with EtOAc
(4.times.75 mL) and the filtrate was concentrated.
[0145] CH.sub.2Cl.sub.2 rinsings gave a pale-yellow solid, 1.7 g,
.sup.1H NMR was consistent with literature values; EtOAc rinsings
gave an off-white solid, 1.0 g, .sup.1H NMR consistent with
literature values; total recover 2.7 g, 75% yield.
(ACL-G29-17)
[0146] The diester (16.4 g, 6.5 mmol) was stirred in anhydrous
hexanes (500 mL) under N.sub.2 and cooled to -78.degree. C. A
solution of DIBAL-H (45 mL, 253 mmol) in hexanes (150 mL) was added
over a 1 h period. Allowed to warm to -30.degree. C. and stirred
overnight (17.5 h total time). A homogeneous mixture of
H.sub.2O/silica gel (12.3 g/43.7 g) was added and the mixture was
manually swirled over a 45 min period. Added K.sub.2CO.sub.3 (15.5
g) and MgSO.sub.4 (23.5 g). Swirled over another 30 min period.
Filtered on a sintered glass funnel, rinsed with methylene chloride
(ppt formed, presumably caused by evaporative cooling) and the
filtrate was concentrated. The solids were rinsed with several
times with EtOAc (approximately 100 mL portions, 2 L total volume)
and pooled with the original filtrate. Concentrated to give a
yellow solid, 8.9 g, 81% crude yield. .sup.1H NMR spectrum was
consistent with the desired product.
2,7-Dimethylocta-2,4,6-triene-1,8-dial (14).sub.2 (ACL-G29-12)
[0147] A slurry of MnO.sub.2 (7.80 g, 90 mmol) was cooled in an ice
bath under N.sub.2. A solution of diol 13 (0.14 g, 0.8 mmol) was
added via pipette as an acetone solution (5.0 mL). An additional
2.0 mL of acetone was used to rinse the flask and complete the
transfer. The ice bath was allowed to melt overnight as the
reaction mixture was stirred. Solids were removed by filtration
through Hyflo and concentrated to give a yellow solid. The material
was dissolved in 10% Et.sub.2O/hexanes with a minimal amount of
CHCl.sub.3 and applied to a column of silica gel (30.times.190 mm)
eluting with 10% Et2O/hexanes. The product could be followed as a
yellow band as it eluted, 14 was isolated as a light yellow solid
37 mg, 26% yield. .sup.1H NMR spectrum was consistent literature
values.
(ACL-G29-16)
[0148] A solution of the diol 13 (2.70 g, 16 mmol) in acetone (500
mL) was cooled in an ice bath under N.sub.2. MnO.sub.2 (60.0 g,
0.69 mol) was added in portions over a 20 min period. The ice bath
was allowed to melt as the reaction mixture was stirred overnight.
The reaction mixture was filtered through Hyflo and the filtrate
was concentrated to give a yellow solid, 1.6 g, 61% crude yield.
.sup.1H NMR was consistent with the literature values. The crude
yellow solid was dissolved in methylene chloride (along with a
small amount of 10% Et.sub.2O in hexanes was added) and charged to
a 4.times.7.5 cm Biotage silica gel column. Eluted initially with
10% ether in hexanes (1 L), then increased polarity to 15%
Et.sub.2O (1 L) and 20% Et.sub.2O (0.5 L). Recovered a yellow solid
1.0 g, 38% yield. .sup.1H NMR spectrum consistent with desired
product.
(ACL-G29-21)
[0149] A solution of the diol (9.31 g, 60 mmol) in acetone (500 mL)
was cooled in an ice bath under N.sub.2. MnO.sub.2 (100 g, 1.15
mol) was added and the mixture was stirred as the ice bath was
allowed to melt overnight. Checked by IR after 24 h, significant
amount of product had formed, but still quite a bit of alcohol
present. Added an additional 50 g of oxidant and continued stirring
for another overnight period. A portion of the reaction mixture was
filtered and checked by .sup.1H NMR, the reaction appeared complete
based on the consumption of starting material. The rest of the
reaction mixture was filtered through a pad of Hyflo and thoroughly
rinsed with acetone. Concentrated to give a dark yellow solid.
Azeotroped once with 40 mL benzene then dried in vacuo at
40.degree. C. for 5 h, then at r.t. overnight. Recovered 5.28 g,
58% yield. .sup.1H NMR and IR spectra were consistent for the
desired product.
Methyl Tiglate (16)
[0150] In a 2L 3-neck flask fitted with an overhead stirrer,
condenser and thermometer, a solution of tiglic acid 15 (89.8 g;
0.9 mol) and 5 mL concentrated sulfuric acid (0.09 mol) in 900 mL
methanol was heated at reflux for 20 hrs. The solution was cooled
to 25.degree. C. and the excess methanol was stripped at 30.degree.
C. and 27 in Hg vacuum on a rotary evaporator. GLC analysis of the
recovered methanol distillate showed product in the overheads. The
resulting two-phase, light brown concentrate was taken up in 500 ml
ethyl ether and washed successively with 250 mL water, 250 mL 10%
aqueous sodium bicarbonate and 250 mL saturated brine. The ether
solution was dried over anhydrous potassium carbonate, filtered and
stripped on the rotary evaporator at 25.degree. C. and 17 in Hg
vacuum to give crude methyl tiglate as a near colorless oil; 43.6 g
(42% yield). GLC analysis showed one major volatile product with a
retention time of 2.7 min compared to 3.8 min for the starting
tiglic acid. Proton NMR in CDCl.sub.3 showed the expected signals
with some trace ethyl ether contamination: 1.79 ppm (d, 3H), 1.83
(s, 3H), 3.73 (s, 3H), 6.86 (q, 6.6 Hz). IR (neat on KBr): ester
carbonyl at 1718 cm.sup.-1. This oil was used as is in the next
step.
Methyl .gamma.-Bromotiglate (17).sub.5
[0151] In a IL 4-neck flask fitted with an overhead stirrer, a
thermometer and a condenser, a stirred mixture of the crude methyl
tiglate (43.6 g; 0.38 mol), N-bromosuccinimide (68 g; 0.38 mol) and
70% benzoyl peroxide (5.34 g; 0.015 mol) in 500 mL carbon
tetrachloride was heated at reflux for two hours. After cooling to
20.degree. C., the insoluble succinimide (38.1 g 100% recovery) was
suction filtered off. The filtrate was washed three times with 250
mL water, dried over MgSO.sub.4 and then stripped on a rotary
evaporator at 25.degree. C. and 26 in Hg vacuum to give a yellow
oil; 78.8 g. Proton NMR of this oil in CDCl.sub.3 gave a complex
spectrum. The methylene protons for the desired y-bromo ester were
assigned to the doublet centered at 4.04 ppm (8.6 Hz), while the
same protons for the a-bromo isomer were ascribed to the singlet at
4.24 ppm. Proton integration of these signals and the methyl
multiplet from 1.6 to 2.0 ppm suggested the following composition
(mole %):
1 .gamma.-bromo ester: 59% .alpha.-bromo ester: 26% starting
material: 15%
[0152] This crude oil was used in the next step without any further
purification.
[0153] This reaction was also run on a 0.05 mole scale using only
0.87 equivalents of N-bromosuccinimide under otherwise identical
conditions. The composition of this crude oil was estimated based
on its proton NMR spectrum as 52% .gamma.-bromo ester, 24%
.alpha.-bromo ester and 23% unreacted methyl tiglate. GLC analysis
of this oil was slightly more complicated showing other minor
components.
Triphenylphosphonium Salt of Methyl .gamma.-Bromotiglate
(19).sub.6
[0154] In a 2L 4-neck flask fitted with a thermometer, a 100 mL
constant pressure addition funnel and a condenser connected to a
static nitrogen system, a stirred solution of the crude methyl
.gamma.-bromotiglate (78.8 g) in 350 ml benzene was treated
dropwise with a soluton of triphenylphosphine (95 g; 0.36 mol) in
350 mL benzene over a period of 1.75 hrs. The temperature of the
mixture exothermed slightly from 24 to 27.degree. C. under
otherwise ambient conditions. After the addition, the reaction was
stirred vigorously overnight to afford a slurry of white solid
containing a yellowish gum that adhered to the walls of the flask.
The white solid was suction filtered onto a sintered glass funnel
without disturbing the yellowish gum. The flask was washed twice
with 100 mL benzene and poured onto the filter. The filter cake was
washed with 50 mL benzene and then twice with 50 mL hexane. The wet
cake was dried in a vacuum oven at ambient temperature for 5.5
hours. The dried white powder [93 g; mp=125.degree. C. dec)] was
dissolved in 150 mL acetonitrile with heat to give a clear yellow
solution. Ethyl acetate (300 mL) was added to this hot solution and
the product started to crystallized after adding about 100 mL ethyl
acetate. The flask was stored in the refrigerator overnight. The
product was suction filtered and washed with a minimum amount of
1:2 acetonitrile and ethyl acetate; 45.0 g. mp=187-190.degree. C.
(dec). lit mp=183.degree. C. (dec).
[0155] The gummy solids in the reaction flask were recrystallized
from 10 mL acetonitrile and 20 mL ethyl acetate. Also, additional
solids precipitated overnight from the benzene mother liquor. These
solids were filtered and recrystallized in the same manner. Both
samples were refrigerated for 2 hours and suction filtered to give
additonal product; 13.3 g.
[0156] The benzene filtrate was stripped on a rotary evaporator and
the yellow oil taken up in 10 mL acetonitrile and precipitated with
20 mL ethyl acetate. The slurry was stored in the refrigerator
overnight to give additional product as a white solid; 4.6 g. m.p.
185-187.degree. C. (dec). Total yield of the desired phosphonium
salt as a white solid was 62.9 g or 36.2% yield based on the crude
methyl tiglate. Proton NMR (CDCl.sub.3, TMS) ppm 1.55 (d, 4 Hz,
3H), 3.57 (s, 3H), 4.9 (dd, 15.8 & 7.9 Hz, 2H), 6.55 (broad q,
6.6-7.9 Hz, 1H), 7.4-7.9 (m, 15H). Proton-decoupled Phosphorus NMR
(CDCl.sub.3, 5% aq H.sub.3PO.sub.4 coaxial external standard) 22.08
ppm. Partial Carbon NMR (CDCl.sub.3): CO.sub.2CH.sub.3, (166.6 ppm,
d, JCP=3 Hz), olefinic CH (117.5 ppm, d, JCP=86.1 Hz), CO.sub.2CH3,
(52.0 ppm), Ph.sub.3P--CH.sub.2 (25.4 ppm, d, J.sub.CP=50.6 Hz) and
CH.sub.3 (13.4 ppm, d, J.sub.CP=2.4 Hz). Partial IR (KBr pellet):
ester carbonyl at 1711 cm.sup.-1.
(3-Carbomethoxy-2-buten-1-ylidene)triphenylphosphorane
(20).sub.6
[0157] In a 5L 5-neck flask fitted with an overhead stirrer, an
addition funnel and a thermometer, a solution of sodium hydroxide
(5.12 g; 0.128 mol) in 250 ml water was added dropwise to a
vigourously stirred solution of the triphenylphosphonium salt of
methyl .gamma.-bromotiglate (58.3 g; 0.128 mol) in 2,500 mL water
over a period of 41 minutes at 25.degree. C. The yellow slurry was
stirred for 10 minutes at room temperature and then suction
filtered. The filter cake was washed with 1,800 mL water and then
thoroughly dried on the filter with a nitrogen blanket. The yellow
solid was then dried overnight in a vacuum desiccator over
P.sub.2O.sub.5 at room temperature and 27" Hg vacuum; 35.3 g (73.7%
yield). mp=145-150.degree. C. lit mp=145-165.degree. C.
Proton-decoupled phosphorus NMR in CDCl.sub.3 showed two peaks at
17.1 ppm and 21.1 ppm in a ratio of 93:7. Proton NMR (CDCl.sub.3,
TMS) ppm 1.89 (s, 3H), 3.58 (s, 3H), 7.3-7.8 (m, 17H). A small but
detectable singlet at 1.74 ppm was also apparent in this spectrum
which was attributed to the impurity. This solid was used without
further purification in the next step.
Dimethyl Crocetinate (21).sub.6 (ACL-G29-18)
[0158] The dialdehyde 14 (0.48 g, 2.9 mmol) was added to a 100 mL
round bottom flask. Benzene (20 mL) was added and the solids were
dissolved with magnetic stirring. The ylide was added, an
additional 10 mL benzene was used to wash the compound into the
flask. Warmed to a vigorous reflux for 6 h. The reaction mixture
was allowed to cool overnight. Contrary to literature reports, a
very small amount of solid had formed. The reaction mixture was
concentrated, the residue was taken up in MeOH (30 mL) and boiled
for 30 min. Upon cooling to ambient temperature, the solids were
collected by vacuum filtration. An NMR sample was prepared by
dissolving 20 mg into 0.5 mL CDCl.sub.3, somewhat surprisingly,
this required warming with a heatgun to dissolve completely.
.sup.1H NMR spectrum was recorded and found to be consistent with
the desired product. The remaining material was dissolved in hot
benzene, filtered, the filtrate was concentrated, taken up in MeOH,
cooled in an ice bath and solids red solids were collected, 334 mg,
33% yield. This material did not appear to be any more soluble than
the material which was originally isolated.
(ACL-G29-18A)
[0159] Dialdehyde 14 (5.78 g, 35 mmol) was dissolved in benzene
(300 mL) under N.sub.2. Ylide 20 (35.3 g, 94 mmol) was added and
the mixture was warmed to reflux for 6 h forming a dark red
solution. After allowing the reaction mixture to cool overnight,
red solids were collected by vacuum filtration and rinsed with
methanol. Transferred to a 500 mL RBF and refluxed with
approximately 65 mL methanol for 30 min. Cooled and collected a red
solid. Rinsed with cold methanol and dried in vacuo to give 21 as a
red solid, 3.00 g. .sup.1H NMR and IR spectra were consistent with
the desired product.
[0160] The original filtrate (from the reaction mixture) was
concentrated on a rotary evaporator and the dark residue was taken
up in 100 mL methanol and refluxed for 40 min. Cooled in an ice
bath and collected by vacuum filtration a red solid. Rinsed with
cold methanol and dried in vacuo to give 21 as a red solid, 1.31 g.
.sup.1H NMR spectrum was consistent with the desired product.
[0161] The filtrates were pooled, concentrated and taken up in 75
mL methanol and allowed to sit overnight at r.t. A red solid was
recovered by vacuum filtration: 0.38 g. .sup.1H NMR spectrum was
consistent with the desired product.
[0162] More solids had formed in the filtrate. Isolated by vacuum
filtration to give a red solid, 0.127 g. IR consistent with above.
Total recovery: 4.89 g, 39% yield.
Saponification Attempt with THF/NaOH (ACL-G29-19)
[0163] A stirred suspension of diester 21 (100 mg, 0.28 mmol) in
THF (2 mL) and 1N NaOH (0.56 mL, 2 eq) was added. Stirred at r.t.
overnight. TLC showed only starting material. Warmed to reflux, no
change after several hours. Added THF (6 mL) in an attempt to
dissolve more of the solids, but it didn't seem to matter.
Continued refluxing overnight. Added more THF (about 6 mL, TLC
showed only starting material), and refluxed for another overnight
period. Concentrated and check by .sup.1H NMR--only starting
material (based on integration of the methyls and methyl esters).
Dissolved in pyridine (10 mL) while warmed on a heating mantle.
Added 2.5 N NaOH (1.0 mL). The dark orange solution turned deep red
after several minutes. The heating mantle was removed, solids began
forming, mantle reapplied for 30 min, then stirred at r.t.
overnight. Concentrated on high vacuum. The residue was insoluble
in chloroform, DMSO, pyridine and sparingly soluble in H.sub.2O. An
IR (Nujol mull) showed C.dbd.O absorbance characteristic of the
starting material.
Saponification with 2.5 N NaOH and THF (ACL-G29-20)
[0164] Diester 21 (37 mg, 0.10 mmol) was weighed into a flask and
stirred in diethyl ether (4 mL). The solvent took on an orange
color, but solids were still present. Added 1 mL of 2.5 N NaOH and
warmed to reflux. After half an hour, most of the ether had
evaporated. This was replaced with THF (3 mL) and refluxing was
continued for several hours. Solid were collected by vacuum
filtration, rinsed with deionized water then dried in a vacuum
oven. IR showed only starting material.
Saponification with 40% NaOH (1) (ACL-G29-22)
[0165] Diester 21 (32 mg, 8.9 mmol) was weighed into a flask and
stirred in methanol (1.5 mL). The solvent took on an orange/red
color, but solids were still present. Added 1.5 mL of 40% NaOH and
warmed to reflux for 17 h. After cooling to r.t., orange solids
were collected by vacuum filtration and rinsed with deionized
water. Dried in vacuo at 40.degree. C. to give 1 as an orange
powder 21 mg, 59%. IR (KBr pellet) 3412, 1544, 1402 cm.sup.-1, the
compound is probably hygroscopic, upfield carbonyl shift is
consistent with conjugation.
(ACL-G29-22A)
[0166] Repeated with 35 mg of diester 1 refluxing for 15 h. The
reaction mixture was cooled in an ice bath, collected by vacuum
filtration and washed with cold deionized water. Dried in vacuo at
40.degree. C. Recovered 1 as an orange solid 25.5 mg, 65%.
(ACL-G29-23)
[0167] Diester 21 (0.48 g, 1.3 mmol) was taken up in methanol (15.0
mL) and 40% sodium hydroxide (15.0 mL) and warmed to reflux. The
heterogeneous red mixture turned orange after about 2 h. Heating
was discontinued after 6 h and the mixture was allowed to cool
overnight. An orange solid was collected by vacuum filtration and
washed with cold deionized water. Drying in vacuo gave a friable
orange solid, 0.36 g, 68% yield.
(AC L-G29-24)
[0168] Diester 21 (1.10 g, 3.1 mmol) was placed in a 100 mL
recovery flask and heated to reflux in methanol (20 mL) and 40%
NaOH (20 mL) for 12 h. After cooling in an ice bath, an orange
solid was collected by vacuum filtration and rinsed with deionized
water. Drying in vacuo gave 1.4 g, 100%. Anal Calcd for
C.sub.20H.sub.220.sub.4Na.sub.2-0.4H.sub.2O: C, 63.29; H, 6.05; Na,
12.11; H.sub.2O, 1.90. Found: C, 63.41; H, 6.26; Na, 11.75;
H.sub.2O, 1.93.
(ACL-G29-25)
[0169] Diester 21 (3.00 g, 8.4 mmol) was refluxed in methanol (80
mL) and 40% NaOH (60 mL) for 12 h. The product was isolated as an
orange solid as described above 2.7 g, 80%. Anal Calcd for
C.sub.20H.sub.220.sub.4Na.sub.- 2-0.4H.sub.2O: C, 63.29; H, 6.05;
Na, 12.11; H.sub.2O, 1.90. Found: C, 63.20; H, 6.00; Na, 11.93;
H.sub.2O, 1.81.5amples ACL-G29-23,-24 and -25 were ground on an
agate mortar and combined as ACL-G29-A.
REFERENCES
[0170] 1. E. Buchta and F. Andree Naturwiss. 1959, 46, 74.
[0171] 2. F. J. H. M. Jansen, M. Kwestro, D. Schmitt, J. Lugtenburg
Recl. Trav. Chim. Pays-Bas 1994, 113, 552.
[0172] 3. R. Gree, H. Tourbah, R. Carrie Tetrahedron Letters 1986,
27, 4983.
[0173] 4. G. M. Coppola Syn. Commun. 1984, 1021.
[0174] 5. D. S. Letham and H. Young Phytochemistry 1971, 10,
2077.
[0175] 6. E. Buchta and F. Andree Chem. Ber. 1960, 93, 1349.
Example 2
[0176] Synthesis of Trans Potassium Norbixinate 14
[0177] Trans potassium norbixinate is synthesized by coupling a
symmetrical C20 dialdehyde containing conjugated carbon-carbon
double bonds with
[1-(ethoxycarbonyl)methylidene]triphenylphosphorane. The
preparation of this compound is similar to that listed previously
for trans sodium crocetinate, except that the furan starting
material is replaced with the appropriate ringed structure. This
product is then saponified using a solution of KOH/methanol.
Example 3
[0178] Synthesis of a Longer BTCS 15
[0179] The above compound is synthesized by adding a symmetrical
C.sub.10 dialdehyde containing conjugated carbon-carbon double
bonds to an excess of
[3-carbomethoxy-2-buten-1-ylidene]triphenylphosphorane. The
preparation of this compound is similar to that listed previously
for trans sodium crocetinate, except that the furan starting
material is replaced with the appropriate ringed structure. The
trans 40-carbon product is then isolated using a procedure such as
chromatography. This product is then saponified using a solution of
NaOH/methanol.
Example 4
[0180] TSC by Inhalation
[0181] TSC has been given to rats via an inhalation route. Ten rats
were given TSC directly into the lungs. This was done by inserting
a tube into the trachea, and nebulizing 0.2 ml of TSC solution (TSC
dissolved in dilute sodium carbonate solution) with about 3 to 6
mls of air. For all dosages studied (0.5-2 mg/kg), about 20% of the
drug was present in the blood stream within one minute after it was
given. For dosages of 0.8-1.6 mg/kg the drug was present in the
blood stream for a period of at least two hours.
Example 5
[0182] Improved Synthesis Method
[0183] Prep of Tetraethyl 2-Butenyl-1,4-bisphosphonate 16
[0184] A 250 mL 3-neck flask was equipped with a Teflon-coated
thermocouple, a 60 mL constant pressure addition funnel and a
simple distillation head. Under a nitrogen atmosphere, neat
triethyl phosphite (59 mL; 0.344 mol) was heated with a heating
mantle controlled with a JKem controller at 140.degree. C. A
solution of trans-1,4-dichloro-2-bute- ne (26.9 g; 0.215 mol) and
triethyl phosphite (35 mL; 0.204 mol) was added dropwise at
134-144.degree. C. over a period of 93 minutes. The clear solution
was then kept at 140.degree. C. under nitrogen. After 37 minutes,
gas chromatography of an aliquot (1 drop) in 1 mL of ethyl acetate
showed desired product, intermediate product and the two starting
materials.
[0185] After 15.5 hrs at 140.degree. C., gas chromatography of an
aliquot (1 drop in 0.5 mL EtOAc) showed the desired product with no
detectable starting dichloride or intermediate product. After 16
hrs, the faint yellow solution was cooled to room temperature under
nitrogen. The faint yellow oil was distilled in a Kugelrohr with a
two-bulb receiver and the further bulb cooled in a dry ice-acetone
bath at 25-100.degree. C. and 0.1-0.2 torr to give a colorless oil
(14.8 g) as a forecut. Gas chromatography showed only product in
the Kugelrohr pot. This light amber oil was distilled in a
Kugelrohr at 140.degree. C. and 0.1-0.15 torr to give distillate as
a colorless oil; 66.45 g (94.1% yield). Gas chromatography showed
only one volatile component. GC-MS analysis showed that this
component was the desired product, giving a small molecular ion at
328 m/z and a base ion at 191 m/z (loss of PO.sub.3Et.sub.2).
Proton NMR was consistent with the desired product. Carbon NMR also
was consistent with the desired bis(phosphonate diester), showing
only long range (W-coupling) and normal carbon-phosphorus coupling
to the allylic carbon.
[0186] Pot residue--light yellow oil -0.8 g.
Prep of 1,1,8,8-Tetramethyoxy-2,7-dimethyl-2,4,6-ocatriene
[0187] 17
[0188] Under a nitrogen atmosphere, a magnetically stirred mixture
of tetraethyl trans-2-butenyl-1,4-bisphosphonate (3.3 g; 10.0
mmol), pyruvic aldehyde dimethyl acetal (2.6 mL; 21.5 mmol) in 10
mL toluene and 10 mL cyclohexane was treated successively with
anhydrous potassium carbonate (10.2 g; 73.8 mmol) and powdered
sodium hydroxide (1.25 g; 31.2 mmol). The solution turned yellow
immediately. The resulting slurry was stirred at ambient
temperature under nitrogen. The reaction slowly exothermed,
reaching a maximum of 38.degree. C. after about 25 minutes. Also, a
gummy precipitated formed, which negatively impacted magnetic
stirring. After 2.5 hrs, gas chromatography of an aliquot of the
yellow-orange solution (1 drop in 0.5 mL toluene) showed the two
starting materials and 3 other new components.
[0189] After 16.75 hrs at ambient temperature, gas chromatography
of an aliquot of the orange solution (1 drop in 0.5 mL toluene)
showed only a small amount of the starting bis(phosphonate
diester). The resulting orange mixture with a gummy mass (unable to
stir) was cooled in an ice bath and quenched with 100 mL 10%
aqueous NaCl. The solids were dissolved in this aqueous solution by
working with a spatula. The mixture was then extracted with 200 mL
1:1 ether:hexane. The organic layer was washed with 10% aqueous
NaCl (200 mL) and then saturated brine (100 mL). The colorless
organic layer was dried over Na.sub.2SO.sub.4. Gas chromatography
showed three major components and no detectable starting
bis(phosphonate diester). The thin layer chromatogram showed two
major spots and one minor spot. The Na.sub.2SO.sub.4 was suction
filtered off and washed with ether. The filtrate was concentrated
on a rotary evaporator at 35.degree. C. to give a colorless oil;
1.8 g. GC-MS Analysis showed that the three major volatile
components were the isomeric products, giving molecular ions at 256
m/z and base ions at 75 m/z [(MeO).sub.2CH.sup.+]. Proton NMR also
was consistent with a mixture of isomeric products along with other
unidentified impurites. Yield of crude product=70.3%.
Prep of 1,1,8,8-Tetramethyoxy-2,7-dimethyl-2,4,6-ocatriene
[0190] 18
[0191] A mechanically stirred mixture of tetraethyl
trans-2-butenyl-1,4-bisphosphonate (63.2 g; 0.19 mol), pyruvic
aldehyde dimethyl acetal (50 mL; 0.41 mol) in 200 mL toluene and
200 mL cyclohexane was treated successively with anhydrous
potassium carbonate (196 g; 1.42 mol) and powdered sodium hydroxide
(24.0 g; 0.60 mol). The solution turned yellow immediately. The
resulting slurry was stirred at ambient temperature under nitrogen.
The reaction exothermed to 61.degree. C. after about 11 minutes and
the stirred mixture was cooled in an ice bath to drop the
temperature to 35.degree. C. After 4.7 hrs at 29-35.degree. C., gas
chromatography of an aliquot (3 drops in 0.5 mL toluene) showed no
starting bis(phosphonate). After 5 hrs, the mixture was cooled in
an ice bath to 13.degree. C. and 10% aqueous sodium chloride (400
mL) was added as the temperature rose to 30.degree. C. More 10%
aqueous sodium chloride (1,500 mL) was added and the mixture was
extracted with 3,000 mL 1:1 ether:hexane. The tinted yellow organic
layer was washed with 10% aqueous sodium chloride (2.times.1,000
mL) and then with saturated brine (1,000 mL). The tinted yellow
organic layer was dried over Na.sub.2SO.sub.4, filtered and
concentrated on a rotary evaporator at 30.degree. C. to give a
light yellow oil; 43.4 g. Gas chromatography showed three major
components comprising 89% of the mixture with no detectable
starting bis(phosphonate). TLC analysis showed one major and 3
minor components.
[0192] Proton NMR showed isomeric product plus toluene. The oil was
evaporated further on a Kugelrohr at 50.degree. C. and 0.2 torr for
30 minutes; 31.9 g. Proton NMR showed isomeric bis(acetal) product
with no detectable toluene.
[0193] Yield=65.5%
Prep of 2,7-Dimethyl-2,4,6-ocatrienedial at Higher Payload
[0194] 19
[0195] Under a nitrogen atmosphere, a magnetically stirred solution
of crude 1,1,8,8-tetramethyoxy-2,7-dimethyl-2,4,6-ocatriene isomers
(31.9 g; 124.4 mmol) in tetrahydrofuran (160 mL), water (80 mL) and
glacial acetic acid (320 mL) was heated at 45.degree. C. with a
heating mantle controlled with a JKem controller via a
Teflon-coated thermocouple (9:03 am). After 30 minutes, the mixture
exothermed to a maximum of 54.degree. C. and then returned to the
45.degree. C. setpoint. Gas chromatography of an aliquot (3 drops
in 0.5 mL THF) after 3 hours showed some residual starting
material, two major and one minor product. The yellow reaction
solution was cooled in an ice bath to 21.degree. C. and then
diluted with 4:1 ether:dichloromethane (2,000 mL). This solution
was then washed successively with 20% aqueous NaCl (2,000
mL.times.2), 4:1 20% aq NaCl: 1M aqueous NaOH (2,000
mL.times.3).sup.1 and 20% aq NaCl (1,000 mL.times.2). The yellow
organic layer was dried over MgSO.sub.4, filtered and concentrated
on a rotary evaporator to give a yellow solid; 18.9 g. Gas
chromatography showed one major and one minor component starting
bis(acetal). TLC analysis showed one major spot and several minor,
more polar impurities. This solid was dissolved in 250 mL refluxing
methanol, cooled to room temperature and then in an ice bath for 1
hr. The slurry was suction filtered to give a yellow fluffy
needles; 14.15 g. Gas chromatography showed 95:5 mixture of
isomeric dialdehydes. This solid was recrystallized again with 200
mL refluxing methanol, cooled to room temperature and then in the
refrigerator overnight. .sup.1The first two washes apparently
removed acetic acid as evident by neutral pH. The third wash turned
red and was still basic, suggesting removal of by product.
[0196] The slurry was suction filtered and washed with
freezer-chilled methanol to give yellow needles; 11.2 g. Gas
chromatography showed 97:3 mixture of isomeric dialdehydes. TLC
analysis showed one spot. The needles were dried in a vacuum oven
at 45.degree. C. for 160 minutes until constant weight; 10.75 g.
uncorrected mp=154-156.degree. C. lit.sup.2 mp=161-162.degree. C.
Proton NMR and Carbon NMR were consistent with the desired
symmetrical dialdehyde. .sup.2 Dictionary of Organic Compounds.
Verson 10:2, Sept, 2002.
[0197] The two methanol filtrates from the recrystallizations were
combined. The thin layer chromatogram showed product plus other
impurities. The filtrates were concentrated and various crops
collected as shown below.
2 Crop Appearance Amt (g) Isomeric Ratio 2 yellow powder 1.4 80:20
3 yellow needles 2.6 75:25 4 yellow solid 4.45 46:30
[0198] Crop 2 & 3: These combined crops were dissolved in 20 mL
refluxing ethyl acetate, cooled to room temperature and then in the
freezer for 1 hr. The slurry was suction filtered and washed with
freezer-chilled ethyl acetate to give yellow needles; 1.95 g. Gas
chromatography showed 86:14 mixture of isomers. This solid was
recrystallized again in ethyl acetate (10 mL) to give yellow
needles; 1.55 g. Gas chromatography showed 92:8 ratio of isomers. A
third recrystallization from ethyl acetate (10 mL) afforded yellow
needles; 1.25 g. mp=152-154.degree. C. Gas chromatography showed
96:4 isomer ratio. Proton NMRconfirmed as the desired dialdehyde.
GC-MS analysis was consistent with the desired dialdehdye, showing
a prominent M.sup.+ ion at 164 m/z and a base ion at 91 m/z.
[0199] The ethyl acetate filtrate was combined with the yellow
solid from the methanol filtrate (crop 4) and concentrated on a
rotary evaporator to give a yellow solid; 6.0 g. Gas chromatography
showed a 53:34 mixture of the two isomers along with other
impurities.
[0200] The solid was dissolved in 100 mL dichloromethane and
Davisil grade 643 silica gel (33.5 g) was added. The mixture was
stripped on a rotary evaporator at 35.degree. C. The silica gel
with adsorbed material was then added to the sample introduction
module for the Biotage system, which already contained a plug of
glass wool and a layer of sand. The silica gel was then topped with
filter paper. The Biotage 75S column was previously wetted with the
solvent mixture with a radial compression of 35 psi and solvent
pressure of 20 psi. The column was eluted with 85:15 hexane:ethyl
acetate (6,000 mL). A void volume of 1,000 mL including the prewet
stage was taken. Fractions of 250 mL were collected and combined
based on thin layer chromatogram analysis. These fractions were
concentrated on a rotary evaporator at 35.degree. C. as shown
below.
3 Fraction Content Appearance Amt (g) Comment 1 blank 2-3 A 4 tr A
5-10 B yellow solid 3.9 Product Cut 11-18 tr B or tr C No evidence
of close eluting impurity 19-20 tr B or C & D
[0201] Fractions 5-10: The yellow solid was slurried in hexane and
suction filtered to give a bright yellow solid; 2.5 g. Gas
chromatography showed an mixture of dialdehyde isomers in a ratio
of 67:33.
[0202] Total yield of 96-97% E,E,E-dialdehyde=10.75+1.25=12.0 g
(58.8% yield).
[0203] Isomerization of 2,7-Dimethyl-2,4,6-ocatrienedial with
para-Toluenesulfinic Acid 20
[0204] Under a nitrogen atmosphere, the 2:1 isomeric mixture of
2,7-dimethyl-2,4,6-ocatrienedial and its off-isomer (2.5 g; 15.2
mmol) and 4-toluenesulfinic acid (0.35 g; 2.2 mmol) and 50 mL
anhydrous 1,4-dioxane was heated at reflux for 15 minutes. An
aliquot (7 drops) was diluted in 0.5 mL 4:1 ether:dichloromethane
and dried over K.sub.2CO.sub.3. Gas chromatography showed a 91:9
mixture of desired isomer and off-isomer.
[0205] After cooling overnight at room temperature, the resulting
slurry was dissolved in 100 mL 4:1 ether:dichloromethane and washed
successively with water (50 mL.times.3), 0.2M aqueous NaOH (50 mL),
water (50 mL.times.2) and saturated brine (50 mL.times.3). After
separation of the layers, the remaining rag layer was dissolved in
dichloromethane. The combined organic layers were dried over
MgSO.sub.4, filtered and concentrated on a rotary evaporator at
40.degree. C. to give an orange solid; 2.2 g. Gas chromatography
showed 93:7 ratio of desired dialdehyde to off-isomer. This solid
was slurried in hexane and suction filtered to give an orange
solid; 2.15 g. This solid was recrystallized from 20 mL refluxing
ethyl acetate by cooling to 30-40.degree. C. and then in the
freezer for 1 hr. The slurry was suction filtered and washed with
freezer-chilled ethyl acetate to give yellow-orange needles; 1.65
g. mp=158-160.degree. C. lit mp=161-162.degree. C.
[0206] Gas chromatography showed 96:4 ratio of desired dialdehyde
to off-isomer. Proton NMR and Carbon NMRwere consistent with the
desired dialdehdye isomer.
[0207] Yield=66%
[0208] Scaleup Prep of Methyl Tiglate with Thionyl Choloride in
Methanol 21
[0209] A mechanically stirred solution of tiglic acid (397.35 g;
3.97 mol) in 3,000 mL methanol was treated dropwise with neat
thionyl chloride (397 mL; 5.44 mol) over a period of 130 minutes as
the temperature climbed from 14.degree. C. to a maximum of
50.degree. C. after 80 minutes with no external cooling. Gas
chromatography of an aliquot showed complete conversion to the
ester with no detectable tiglic acid. After stirring at ambient
temperature for 1 hr, the solution was distilled at atmospheric
pressure through a silvered, vacuum jacketed Vigreux column (400
mm.times.20 mm). The condensate was collected at mainly
57-61.degree. C. with a pot temperature of 58-63.degree. C.; 630 mL
in 2 hrs. Gas chromatography showed significant methyl ester in the
distillate.
[0210] The Vigreux column was swapped with a less efficient column
(30.times.2 cm w/less indentations) to speed up the rate of
distillation. At a pot temperature of 69-71.degree. C., distillate
was collected with a head temperture of 65-69.degree. C.; 1,300 mL
over 2.25 hrs.
[0211] Gas chromatography showed significant methyl ester in the
distillate. The atmospheric distillation was continued until the
pot temperature reached 87.degree. C., distillate was collected
during this period at a head temperture of 69-83.degree. C.; 975 mL
over 2 hrs. Gas chromatography showed significantly more methyl
ester in the distillate than earlier fractions.
[0212] The yellow two-phase mixture in the pot was extracted with
ether (300 & 200 mL), dried over K.sub.2CO.sub.3, filtered and
concentrated on a rotary evaporator at 25.degree. C. to give an
orange oil; 132.6 g (29.3% yield). Gas chromatography showed
product. Proton NMR and carbon NMR were consistent with the desired
product with trace ethyl ether. Gas chromatography of the ether
condensate showed some methyl ester in the overheads.
[0213] Distillate 3: The third methanol distillate (975 mL) was
concentrated on the rotary evaporator at 25.degree. C. to give a
two phase mixture (100-150 mL). This mixture was extracted with
ether (100 & 50 mL), dried over K.sub.2CO.sub.3.
[0214] Distillate 2: The second methanol distillate (1,300 mL) was
concentrated on the rotary evaporator at 25.degree. C. to give a
two phase mixture (30-50 mL). This mixture was extracted with ether
(2.times.50 mL), dried over K.sub.2CO.sub.3.
[0215] The concentrated ether extracts for distillate 2 and
distillate 3 were combined, suction filtered and concentrated on a
rotary evaporator at 25.degree. C. to give a colorless oil; 77.3
g.
[0216] Proton NMR and carbon NMR matched previous spectra of the
desired methyl ester.
[0217] Total Yield=132.6+77.3=209.9 g (46.3%)
[0218] Alternatively, 1) methyl tiglate is commercially available
from Alfa, Lancaster or Acros. and 2), pilots can be run to make
phosphonium salt via JOC, 64, 8051-8053 (1999).
[0219] Bromination of Methyl Tiglate 22
[0220] A mechanically stirred slurry of methyl tiglate (209.9 g;
1.84 mol) and N-bromosuccinimide (327.5 g; 1.84 mol), 70% benzoyl
peroxide (3.2 g; 0.009 mol) in 2,000 mL carbon tetrachloride was
heated to reflux (78-81.degree. C.) with a IL Kugelrohr bulb
between the 5L reaction flask and the reflux condenser. After 2
hrs, reflux was stopped, the mantle dropped and the stirrer
shutoff. All of the solids floated on the CCl.sub.4 solution,
suggesting succinimide with negligible NBS. The slurry was cooled
in an ice bath to 20.degree. C. and suction filtered to give an
offwhite solid; 180.7 g. No wash. The yellow filtrate was washed
with water (1L.times.3), dried over MgSO.sub.4. Gas chromatography
showed starting methyl tiglate and the two monobromides in 1:2:1
ratio along with other minor components.
[0221] After filtering off the MgSO.sub.4, the light yellow
filtrate was concentrated on a rotary evaporator at 35.degree. C.
to give a light yellow oil; 327.1 g. Proton NMR and gas
chromatography suggested the following composition:
4 Component NMR (mole %) GC (Area %) .gamma.-Bromo 50% 49%
.alpha.-Bromo 26% 21% .alpha.,.gamma.-Dibromo (?) 7% 4% Methyl
Tiglate 6% 10% Other 11% --
[0222] Yield of desired product adjusted for 50% assay=46.0%
[0223] This oil is used as is in the next step.
[0224] Scaleup Reaction of Methyl y-Bromotiglate with
Triphenylphosphine in Acetonitrile with Slightly Higher Payload
23
[0225] Under a nitrogen atmosphere in a 5L, 4-neck flask, the crude
mixture of methyl .gamma.-bromotiglate (322.6 g; 85% allylic
bromide; 1.42 mol) in 1,300 mL anhydrous acetonitrile was stirred
mechanically.
[0226] A solution of triphenylphosphine (387.0 g; 1.48 mol) in
2,000 mL ethyl acetate was added dropwise over a period of 4 hours.
During the addition, the temperature climbed from 22.degree. C. to
a maximum of 30.degree. C. after adding about 40% in the first 75
minutes. After adding 60% of the triphenylphosphine solution over
120 minutes, the solution became cloudy and continued to
precipitate solids through the rest of the addition. After the
addition, the funnel was rinsed with ethyl acetate (600 mL) and
chased into the reaction mixture. The cream slurry was stirrred at
ambient temperature over the weekend.
[0227] The white slurry was suction filtered and the cake was
washed with 2:1 ethyl acetate:acetonitrile (150 mL.times.3). The
white solid (352.55 g) was dried in a vacuum oven at 40.degree. C.
for 4 hrs (constant weight after 2 hrs); 322.55 g.
mp=187-188.degree. C. (dec). lit mp=183.degree. C. (dec). Proton
NMRand Carbon NMR matched previous spectra for the desired
phosphonium salt. LC-MS analysis showed one major component, whose
electrospray mass spectrum in the positive mode was consistent with
the desired phosphonium salt giving a molecular ion at 375 m/z.
Phosphorus NMR showed a single phosphorus signal at 22.0 ppm.
[0228] Yield based on starting methyl
tiglate=100.times.322.55/(455.32.tim-
es.1.84.times.322.6/327.1)=39.0%
[0229] Prep of
(3-Carbomethoxy-2-Z-buten-1-ylidene)triphenylphosphorane 24
[0230] A mechnically stirred slight slurry of
(3-carbomethoxy-2-E-buten-1-- ylidene)triphenylphosphonium bromide
(154.8 g; 0.34 mol) in 3,400 mL deionized water was treated
dropwise with a solution of sodium hydroxide (13.6 g; 0.34 mol) in
500 mL water at 23.degree. C. over a period of 32 minutes with no
obvious exotherm, but immediate precipitation of a bright yellow
solid. After stirring for 15 minutes, the bright yellow slurry was
suction filtered, washed with water (1,500 mL) and sucked dry to
give a canary yellow solid; 151.7 g. This solid was dried in a
vacuum oven at 35-45.degree. C. (3:50 pm) overnight.
[0231] After drying in the vacuum oven at 35-45.degree. C. for 22.5
hrs, a constant weight was obtained; 107.8 g. mp=144-160.degree. C.
lit mp=145-165.degree. C. Proton NMR was similar to the previous
spectrum of the desired ylide considering the differences in NMR
field strength. Carbon NMRshowed the methyl carbon's at 50.2 and
11.8 ppm with a complex aromatic region and no obvious signals for
the olefinic carbons and the ylide carbon.
[0232] Yield=84.7%
[0233] Pilot Prep of Dimethyl Crocetinate 25
[0234] Under a nitrogen atmosphere, a magnetically stirred mixture
of (3-carbomethoxy-2-Z-buten-1-ylidene)triphenylphosphorane (12.8
g; 34.2 mmol) and 2,7-dimethyl-2,4,6-ocatrienedial (2.1 g; 12.8
mmol) in benzene (128 mL) was heated to reflux for 6 hrs using a
timer.
[0235] The resulting slurry was cooled in an ice bath for 40
minutes, suction filtered, washed with benzene and sucked dry to
melt the frozen benzene to give a red solid; 2.1 g. TLC analysis
showed a single, yellow spot. This solid was dried in a vacuum oven
at 40-45.degree. C. for 70 minutes; 1.85 g (40.5% yield).
uncorrected mp=210-213.degree. C. lit.sup.3 mp=214-216.degree. C.
Proton NMR was similar to the previous spectrum of dimethyl
crocetin on 90 MHz instrument. Carbon NMR showed all 11 unique
carbon signals with the correct chemical shift for the desired
dimethyl ester with one minor impurity signal that may be residual
benzene. Electrospray mass spectrum suggested decomposition and
recombination of fragments. .sup.3 E. Buchta & F. Andree, Chem
Ber, 93,1349 (1960).
[0236] TLC analysis showed that the red filtrate contained
additional product, triphenylphosphine oxide and an orange
component with an R.sub.f slightly lower than the isolated solid.
The red filtrate was concentrated on a rotary evaporator at
35.degree. C. to give red solids; 13.2 g. This solid was heated at
reflux in methanol (25 mL). The resulting slurry was then cooled in
an ice bath, suction filtered after 60 minutes and washed with
methanol to give a red solid; 0.6 g. This solid was dried in the
vacuum oven at 45.degree. C. 135 minutes; 0.5 g. mp=203-208.degree.
C. Proton NMR showed desired diester with residual impurities.
Carbon NMR showed only signals for desired product. TLC analysis
showed streaky product spot.
[0237] Filtrate was concentrated and saved.
[0238] Second Prep of Dimethyl Ester of Crocetin 26
[0239] Under a nitrogen atmosphere,
2,7-dimethyl-2,4,6-ocatrienedial (11.95 g; 12.8 mmol) was added in
one portion to a mechanically stirred slurry of
(3-carbomethoxy-2-Z-buten-1-ylidene)triphenylphosphorane (73.0 g;
195.0 mmol) in 400 mL benzene and then chased with 330 mL benzene.
The resulting brown slurry was heated to reflux for 6 hrs using a
timer and cooled to room temperature overnight under nitrogen.
[0240] The resulting slurry was cooled in an ice bath to
6-10.degree. C., suction filtered and washed with benzene (50
mL.times.2) to give a red solid; 10.05 g. TLC analysis showed a
single yellow spot. This solid was dried in a vacuum oven at
40.degree. C. (9:00 am) for 3.5 hrs with no weight loss; 10.05 g
(38.7% yield). mp=211-214.degree. C. lit mp=mp=214-216.degree. C.
Proton NMR and Carbon NMR matched the previous spectra for the
desired dimethyl ester of crocetin.
[0241] The red filtrate was concentrated on a rotary evaporator at
40.degree. C. to give a red solid; 84.4 g. TLC analysis was similar
to the pilot run. This solid was slurried in 165 mL methanol at
reflux with magnetic stirring. The resulting slurry was then cooled
in an ice bath for 2.5 hrs, suction filtered and washed with a
minimal amount of methanol to give an orange paste; 10.5 g. TLC
analysis showed a single, yellow spot. This paste was dried in a
vacuum oven at 45.degree. C. for 190 minutes; 5.6 g.
mp=201-208.degree. C. NMR showed desired diester with unknown
aromatic impurities.
[0242] This impure solid and two other similar solids from earlier
runs totaling 6.5 g were dissolved in refluxing chloroform (75 mL)
and diluted with methanol and cooled in the refrigerator
overnight.
[0243] The slurry was suction filtered and washed with a minimal
amount of methanol to give red crystalline solid; 6.1 g. This solid
was dried in the vacuum oven at 45.degree. C. for 3 hrs until
constant weight; 4.25 g. mp=211-213.degree. C. Proton NMR and
carbon NMR showed other olefinic or aromatic impurities. The solid
was dissolved in refluxing toluene (150 mL) and eventually cooled
in the refrigerator for 130 minutes. The slurry was suction
filtered and washed with tolene to give a red solid; 2.05 g. This
solid was dried in the vacuum oven at 45.degree. C. for 50 minutes
with no weight change; 2.05 g. mp=214-216.degree. C. Proton NMR
showed the desired dimethyl crocetin with some residual toluene and
negligible off-isomer impurities. Carbon NMR showed the desired
dimethyl crocetin with no detectable off-isomer impurities and 2-3
new residual signals that were consistent with toluene.
Yield=45.5%.
[0244] Prep of Disodium Salt of Crocetin 27
[0245] A mechanically stirred slurry of dimethyl crocetin (13.95 g;
39.1 mmol) and 40 wt % aqueous sodium hydroxide (273 mL; 3.915 mol)
and methanol (391 mL) was heated at reflux at 74.degree. C. for 12
hrs using a timer.
[0246] The orange slurry was suction filtered through a Buchner
funnel with filter paper and a sintered glass funnel. Slow
filtration..sup.4 The slurry in the sintered glass funnel was added
to the solids in the Buchner funnel. The orange paste was washed
with water (100 mL.times.3) and then with methanol (50 mL x 3). The
orange paste was dried in a vacuum oven at 45-50.degree. C. .sup.4
Filtered faster through sintered glass until the filter clogged
after drying out. However, water wash unclogged the filter.
[0247] After 21 hrs, the orange clumps weighted 24.25 g. The
material was pulverized with a spatula and dried in the vacuum oven
at 45-50.degree. C.
[0248] After a total of 65.5 hrs of drying, amount of orange powder
was 23.1 g. The infrared spectrum showed extra bands compared to
the reported IR spectrum of TSC, especially large bands at 3424 and
1444 cm.sup.-1. Proton NMR showed no evidence of methyl esters.
However, the integration of the olefinic and methyl regions were
off, possibly due to phasing problems.
[0249] Assuming that the excess weight was due to sodium hydroxide,
the orange solid was stirred magnetically in 400 mL deionized water
for 35 minutes. The slurry was suction filtered and the cake washed
with deionized water (50 mL.times.2) to give an orange paste. This
material was dried in a vacuum oven at 45-50.degree. C. until
constant weight. After about 7 hrs, the solid was crushed and
pulverized and dried further in the vacuum oven at 45.degree. C.
overnight.
[0250] After 21 hrs of drying at 45.degree. C., amount of solid was
13.25 g. After further pulverizing and drying in the vacuum oven at
45.degree. C., amount of solid was 13.15 g. The infrared spectrum
was consistent with the reported IR spectrum. Proton NMR gave a
proton NMR spectrum that was consistent with The disodium salt.
HPLC analysis showed one major component with possibly one minor
impurity. The electrospray negative ion mass spectrum of the major
component was consistent with the desired disodium salt of
crocetin. Carbon NMR showed all ten unique carbon signals for
disodium salt of crocetin, verifying the symmetry of the
molecule.
[0251] The original filtrate of water, sodium hydroxide and
methanol precipitated more solids during the water wash. This
slurry was suction filtered, washed with water to give an orange
paste. This paste was dried in the vacuum oven at 45.degree. C. for
18.5 hrs to give an orange solid; 0.65 g. The spectral data were
consistent with the desired disodium salt of crocetin. This solid
was combined with the first crop.
[0252] Yield=13.15+0.65=13.8 g (94.8%).
[0253] Elemental Analyses of the first crop showed unacceptable
values for the desired product, suggesting sodium hydroxide
contamination of the disodium salt of crocetin.
[0254] Water Wash of Disodium Salt of Crocetin
[0255] The disodium salt of crocetin (13.6 g) was slurried in 150
mL deionized water and stirred magnetically at room temperature for
1 hr. The slurry was suction filtered onto a Buchner funnel. The
orange paste was then washed with water and the pH of the orange
filtrate monitored.
[0256] The orange paste was sucked dry on the filter with a rubber
dam. This paste was dried in a vacuum at 25-55.degree. C. for 5.5
hrs to give a friable orange solid; 11.2 g. This solid was
pulverized, transferred to a bottle and dried in the vacuum oven at
45.degree. C. overnight.
[0257] Amount=11.1 g. Recovery=81.6%. The IR and Proton NMR spectra
matched previous IR and proton NMR spectra of the desired disodium
salt of crocetin. HPLC analysis showed a single component at 420
nm, whose electrospray mass spectrum in the negative ion mode was
consistent with crocetin.
[0258] Carbon NMR showed all ten unique carbon signals with the
correct chemical shifts for the desired disodium salt of crocetin.
Elemental analysis gave acceptable data for the desired
product.
REFERENCES
[0259] 1. Tetrahedron Letters, 27, 4983-4986 (1986).
[0260] 2. F. J. H. M. Jansen, M. Kwestro, D. Schmitt & J.
Lugtenburg, Recl. Trav. Chem. Pays-Bas, 113, 552-562 (1994) and
references cited therein.
[0261] 3. J. H. Babler, U.S. Pat. No. 4,107,030, Apr. 21, 1992.
[0262] 4. T. W. Gibson & P. Strassburger, J. Org. Chem., 41,
791 (1976) & J. M. Snyder & C. R. Scholfield, J. Am. Oil
Chem. Soc., 59, 469 (1982).
Example 6
[0263] Purity Determination of TSC Made According to the Improved
Synthesis Method
[0264] For the TSC material synthesized according to the method of
Example 5, the ratio of the absorbance at 421 nm to the absorbance
at 254 nm was 11.1 using a UV-visible spectrophotometer.
Example 7
[0265] Oral Administration of TSC
[0266] TSC has been shown, in rats, to be absorbed into the blood
stream when administered orally (via a gavage technique). In two
rats, it was found that 1 to 2% of the dosage given was present in
the blood stream at a time of 15 to 30 minutes after being given.
The maximum amount absorbed orally actually occurred earlier than
that time.
[0267] It will be readily apparent to those skilled in the art that
numerous modifications and additions can be made to both the
present compounds and compositions, and the related methods without
departing from the invention disclosed.
* * * * *